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95 rows where devstage_curation = "Gastrula" and experiment.library_selection = "other"

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tissue_curation 4

technology 4

experiment.library_strategy 3

experiment.library_layout 2

experiment.platform 1

experiment.library_source 1

experiment.library_selection 1

  • other · 95 ✖

devstage_curation_coarse 1

devstage_curation 1

  • Gastrula · 95 ✖
Link rowid ▼ run.accession experiment.accession sample.accession study.accession bioproject study.title study.alias study.type study.abstract study.attributes study.PMIDs sample.description sample.title sample.alias sample.centername sample.attributes GEOsample.title GEOsample.dataprocessing GEOsample.source GEOsample.treatmentprotocol GEOsample.extractprotocol GEOsample.growthprotocol GEOsample.characteristics GEOsample.accession experiment.title experiment.alias experiment.library_name experiment.design_description experiment.library_construction_protocol experiment.attributes experiment.library_strategy experiment.library_source experiment.library_selection experiment.library_layout experiment.platform experiment.instrument_model experiment.spot_descriptor experiment.study_ref run.title run.attributes run.filename run.semantic_name run.total_bases run.total_spots run.alias run.read_lengths run.base_counts run.r1_length run.r2_length run.r3_length run.r4_length run.Acount run.Ccount run.Gcount run.Tcount run.Ncount run.experiment run.pool_member submission.accession submission.srasource submission.bioprojectsource seqdetective.n_mates seqdetective.mapping_rate.mate1 seqdetective.mapping_rate.mate2 seqdetective.nofeature_rate.mate1 seqdetective.nofeature_rate.mate2 seqdetective.sparsity.mate1 seqdetective.sparsity.mate2 seqdetective.pos_strand_rate.mate1 seqdetective.pos_strand_rate.mate2 seqdetective.readlen.mate1 seqdetective.readlen.mate2 seqdetective.judgement.mate1 seqdetective.judgement.mate2 seqdetective.judgement.reason platform_family instrument_generation read_bias selection_class prep_kit sc_or_bulk tech_class technology tech_variant submission.bioprojectsource.country earliest_date devstage_curation devstage_curation_coarse tissue_curation tissue_curation_coarse
60 60 DRR032764 DRX029570 DRS049969 DRP003810 PRJDB3785 EXPANDE project DRP003810 Other EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief taking advantages of Illumina sequencing RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.     mRNA extracted from pooled embryos of 100 individuals Dr shield 2 SAMD00028161   sample name:Dr shield 2|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:shield|genotype:wild type|phenotype:wild type|sex:male female and mixed                 Illumina HiSeq 2000 sequencing of SAMD00028161 DRX029570 Dr shield 2 1 Total RNA QIAGEN RNeasy followed by TruSeq   RNA-Seq TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2000 <SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR> DRP003810 Illumina HiSeq 2000 sequencing of SAMD00028161       3644397900.0 36443979.0 DRR032764 0:100 1:0 A:986071173;C:842367218;G:837686080;T:978236607;N:36822 100 0     986071173 842367218 837686080 978236607 36822 DRX029570 DRS049969 DRA003460 UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo 1 0.92419   0.08269   0.75558   0.47863   100   B   usable mapping rate illumina hiseq_era unknown other trueseq bulk unknown unknown   Japan 2017-09-20 Gastrula Embryo Whole Organism All anatomical structures
61 61 DRR032763 DRX029569 DRS049968 DRP003810 PRJDB3785 EXPANDE project DRP003810 Other EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief taking advantages of Illumina sequencing RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.     mRNA extracted from pooled embryos of 100 individuals Dr shield 1 SAMD00028160   sample name:Dr shield 1|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:shield|genotype:wild type|phenotype:wild type|sex:male female and mixed                 Illumina HiSeq 2000 sequencing of SAMD00028160 DRX029569 Dr shield 1 1 Total RNA QIAGEN RNeasy followed by TruSeq   RNA-Seq TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2000 <SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR> DRP003810 Illumina HiSeq 2000 sequencing of SAMD00028160       3834622000.0 38346220.0 DRR032763 0:100 1:0 A:1043352851;C:880011834;G:876775415;T:1034444253;N:37647 100 0     1043352851 880011834 876775415 1034444253 37647 DRX029569 DRS049968 DRA003460 UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo 1 0.92305   0.09126   0.75481   0.47587   100   B   usable mapping rate illumina hiseq_era unknown other trueseq bulk unknown unknown   Japan 2017-09-20 Gastrula Embryo Whole Organism All anatomical structures
69 69 DRR032755 DRX029561 DRS049960 DRP003810 PRJDB3785 EXPANDE project DRP003810 Other EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief taking advantages of Illumina sequencing RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.     mRNA extracted from pooled embryos of 100 individuals Dr 90epiboly 2 SAMD00028152   sample name:Dr 90epiboly 2|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:90epiboly|genotype:wild type|phenotype:wild type|sex:male female and mixed                 Illumina HiSeq 2000 sequencing of SAMD00028152 DRX029561 Dr 90epiboly 2 1 Total RNA QIAGEN RNeasy followed by TruSeq   RNA-Seq TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2000 <SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR> DRP003810 Illumina HiSeq 2000 sequencing of SAMD00028152       3572358600.0 35723586.0 DRR032755 0:100 1:0 A:971653450;C:821326559;G:816855636;T:962477457;N:45498 100 0     971653450 821326559 816855636 962477457 45498 DRX029561 DRS049960 DRA003460 UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo 1 0.92485   0.10642   0.74213   0.47012   100   B   usable mapping rate illumina hiseq_era unknown other trueseq bulk unknown unknown   Japan 2017-09-20 Gastrula Embryo Whole Organism All anatomical structures
70 70 DRR032754 DRX029560 DRS049959 DRP003810 PRJDB3785 EXPANDE project DRP003810 Other EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief taking advantages of Illumina sequencing RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.     mRNA extracted from pooled embryos of 100 individuals Dr 90epiboly 1 SAMD00028151   sample name:Dr 90epiboly 1|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:90epiboly|genotype:wild type|phenotype:wild type|sex:male female and mixed                 Illumina HiSeq 2000 sequencing of SAMD00028151 DRX029560 Dr 90epiboly 1 1 Total RNA QIAGEN RNeasy followed by TruSeq   RNA-Seq TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2000 <SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR> DRP003810 Illumina HiSeq 2000 sequencing of SAMD00028151       3423980500.0 34239805.0 DRR032754 0:100 1:0 A:933088185;C:785251613;G:780911148;T:924686406;N:43148 100 0     933088185 785251613 780911148 924686406 43148 DRX029560 DRS049959 DRA003460 UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo 1 0.92436   0.10881   0.74255   0.47068   100   B   usable mapping rate illumina hiseq_era unknown other trueseq bulk unknown unknown   Japan 2017-09-20 Gastrula Embryo Whole Organism All anatomical structures
73 73 DRR032751 DRX029557 DRS049956 DRP003810 PRJDB3785 EXPANDE project DRP003810 Other EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief taking advantages of Illumina sequencing RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.     mRNA extracted from pooled embryos of 100 individuals Dr 75epiboly 2 SAMD00028148   sample name:Dr 75epiboly 2|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:75epiboly|genotype:wild type|phenotype:wild type|sex:male female and mixed                 Illumina HiSeq 2000 sequencing of SAMD00028148 DRX029557 Dr 75epiboly 2 1 Total RNA QIAGEN RNeasy followed by TruSeq   RNA-Seq TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2000 <SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR> DRP003810 Illumina HiSeq 2000 sequencing of SAMD00028148       3252021500.0 32520215.0 DRR032751 0:100 1:0 A:885527595;C:746750899;G:742907892;T:876794123;N:40991 100 0     885527595 746750899 742907892 876794123 40991 DRX029557 DRS049956 DRA003460 UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo 1 0.92594   0.10181   0.74862   0.47789   100   B   usable mapping rate illumina hiseq_era unknown other trueseq bulk unknown unknown   Japan 2017-09-20 Gastrula Embryo Whole Organism All anatomical structures
74 74 DRR032750 DRX029556 DRS049955 DRP003810 PRJDB3785 EXPANDE project DRP003810 Other EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief taking advantages of Illumina sequencing RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.     mRNA extracted from pooled embryos of 100 individuals Dr 75epiboly 1 SAMD00028147   sample name:Dr 75epiboly 1|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:75epiboly|genotype:wild type|phenotype:wild type|sex:male female and mixed                 Illumina HiSeq 2000 sequencing of SAMD00028147 DRX029556 Dr 75epiboly 1 1 Total RNA QIAGEN RNeasy followed by TruSeq   RNA-Seq TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2000 <SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR> DRP003810 Illumina HiSeq 2000 sequencing of SAMD00028147       3785053700.0 37850537.0 DRR032750 0:100 1:0 A:1029014798;C:870946157;G:867537069;T:1017508684;N:46992 100 0     1029014798 870946157 867537069 1017508684 46992 DRX029556 DRS049955 DRA003460 UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo 1 0.92346   0.10046   0.74921   0.47295   100   B   usable mapping rate illumina hiseq_era unknown other trueseq bulk unknown unknown   Japan 2017-09-20 Gastrula Embryo Whole Organism All anatomical structures
9717 9717 ERR3842002 ERX3854564 ERS4268611 ERP116106 PRJEB33323 Deconstructing the individual steps of vertebrate translation initiation ena-STUDY-Computational Biology Unit-03-07-2019-10:11:34:314-422 Other In eukaryotes the number of ribosomes synthesizing a given protein depends on how many are recruited to its mRNA their success in navigating its five prime untranslated region UTR and whether they recognize its start codon. Initiation of translation is a rate limiting step in protein synthesis and key to gene expression control 1 but despite this centrality it remains poorly understood 2 3. Here we introduce ribosome complex profiling RCP seq to capture the transcriptome wide occupancy of scanning initiating elongating and terminating ribosome complexes in a higher eukaryote. We track scanning and elongating ribosomes across all five prime UTRs in zebrafish which enable us to assess the individual regulatory contributions from the three stages of initiation: ribosome recruitment scanning of the five prime UTR and recognition of the start codon. Our data sheds light on small subunit recruitment to mRNAs presenting evidence for the threading model and demonstrates that sequence features regulate this recruitment. We estimate the processivity of scanning ribosomes as they traverse the five prime UTR and show that the repressive effects of upstream open reading frames depend on the efficiency of both translation initiation and termination. Finally we determine the optimal initiation contexts by directly estimating the conversion of scanning to elongating ribosomes and demonstrate specific regulation of translation initiation at the endoplasmic reticulum. Our results open for the possibility of deconvoluting translation initiation into separate stages and provides the first view of global occupancy of ribosomal small subunits in a vertebrate. ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 07 03     Shield 4Ei SAMEA6504165 Computational Biology Unit ENA FIRST PUBLIC:2020 03 27T17:05:06Z|ENA LAST UPDATE:2020 01 27T16:04:35Z|External Id:SAMEA6504165|INSDC center name:Computational Biology Unit|INSDC first public:2020 03 27T17:05:06Z|INSDC last update:2020 01 27T16:04:35Z|INSDC status:public|Submitter Id:Shield 4Ei|common name:zebrafish|sample name:Shield 4Ei|scientific name:Danio rerio                 NextSeq 500 sequencing ena EXPERIMENT Computational Biology Unit 27 01 2020 17:26:02:557 2 Shield 4Ei LSU OTHER RCP seq   OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   ERP116106 NextSeq 500 sequencing ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2020 02 14     10915827408.0 143629308.0 ena RUN Computational Biology Unit 27 01 2020 17:26:02:557 2 0:76 A:3900515347;C:2409375725;G:3041696977;T:1564127293;N:112066 76       3900515347 2409375725 3041696977 1564127293 112066 ERX3854564 ERS4268611 ERA2359340 Computational Biology Unit|European Nucleotide Archive Computational Biology Unit 1 0.64398   0.40944   0.98817   0.59337   76   B   usable mapping rate illumina nextseq unknown other unknown bulk unknown unknown   Unknown 2019-07-03 Gastrula Embryo Undetermined Embryo Imprecise
9718 9718 ERR3842001 ERX3854563 ERS4268611 ERP116106 PRJEB33323 Deconstructing the individual steps of vertebrate translation initiation ena-STUDY-Computational Biology Unit-03-07-2019-10:11:34:314-422 Other In eukaryotes the number of ribosomes synthesizing a given protein depends on how many are recruited to its mRNA their success in navigating its five prime untranslated region UTR and whether they recognize its start codon. Initiation of translation is a rate limiting step in protein synthesis and key to gene expression control 1 but despite this centrality it remains poorly understood 2 3. Here we introduce ribosome complex profiling RCP seq to capture the transcriptome wide occupancy of scanning initiating elongating and terminating ribosome complexes in a higher eukaryote. We track scanning and elongating ribosomes across all five prime UTRs in zebrafish which enable us to assess the individual regulatory contributions from the three stages of initiation: ribosome recruitment scanning of the five prime UTR and recognition of the start codon. Our data sheds light on small subunit recruitment to mRNAs presenting evidence for the threading model and demonstrates that sequence features regulate this recruitment. We estimate the processivity of scanning ribosomes as they traverse the five prime UTR and show that the repressive effects of upstream open reading frames depend on the efficiency of both translation initiation and termination. Finally we determine the optimal initiation contexts by directly estimating the conversion of scanning to elongating ribosomes and demonstrate specific regulation of translation initiation at the endoplasmic reticulum. Our results open for the possibility of deconvoluting translation initiation into separate stages and provides the first view of global occupancy of ribosomal small subunits in a vertebrate. ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 07 03     Shield 4Ei SAMEA6504165 Computational Biology Unit ENA FIRST PUBLIC:2020 03 27T17:05:06Z|ENA LAST UPDATE:2020 01 27T16:04:35Z|External Id:SAMEA6504165|INSDC center name:Computational Biology Unit|INSDC first public:2020 03 27T17:05:06Z|INSDC last update:2020 01 27T16:04:35Z|INSDC status:public|Submitter Id:Shield 4Ei|common name:zebrafish|sample name:Shield 4Ei|scientific name:Danio rerio                 NextSeq 500 sequencing ena EXPERIMENT Computational Biology Unit 27 01 2020 17:26:02:557 1 Shield 4Ei SSU OTHER RCP seq   OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   ERP116106 NextSeq 500 sequencing ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2020 02 14     7154041880.0 94132130.0 ena RUN Computational Biology Unit 27 01 2020 17:26:02:557 1 0:76 A:2939474250;C:1489083073;G:1922522149;T:802890185;N:72223 76       2939474250 1489083073 1922522149 802890185 72223 ERX3854563 ERS4268611 ERA2359340 Computational Biology Unit|European Nucleotide Archive Computational Biology Unit 1 0.4369   0.25417   0.9867   0.60047   76   B   usable mapping rate illumina nextseq unknown other unknown bulk unknown unknown   Unknown 2019-07-03 Gastrula Embryo Undetermined Embryo Imprecise
9719 9719 ERR3842000 ERX3854562 ERS4268611 ERP116106 PRJEB33323 Deconstructing the individual steps of vertebrate translation initiation ena-STUDY-Computational Biology Unit-03-07-2019-10:11:34:314-422 Other In eukaryotes the number of ribosomes synthesizing a given protein depends on how many are recruited to its mRNA their success in navigating its five prime untranslated region UTR and whether they recognize its start codon. Initiation of translation is a rate limiting step in protein synthesis and key to gene expression control 1 but despite this centrality it remains poorly understood 2 3. Here we introduce ribosome complex profiling RCP seq to capture the transcriptome wide occupancy of scanning initiating elongating and terminating ribosome complexes in a higher eukaryote. We track scanning and elongating ribosomes across all five prime UTRs in zebrafish which enable us to assess the individual regulatory contributions from the three stages of initiation: ribosome recruitment scanning of the five prime UTR and recognition of the start codon. Our data sheds light on small subunit recruitment to mRNAs presenting evidence for the threading model and demonstrates that sequence features regulate this recruitment. We estimate the processivity of scanning ribosomes as they traverse the five prime UTR and show that the repressive effects of upstream open reading frames depend on the efficiency of both translation initiation and termination. Finally we determine the optimal initiation contexts by directly estimating the conversion of scanning to elongating ribosomes and demonstrate specific regulation of translation initiation at the endoplasmic reticulum. Our results open for the possibility of deconvoluting translation initiation into separate stages and provides the first view of global occupancy of ribosomal small subunits in a vertebrate. ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 07 03     Shield 4Ei SAMEA6504165 Computational Biology Unit ENA FIRST PUBLIC:2020 03 27T17:05:06Z|ENA LAST UPDATE:2020 01 27T16:04:35Z|External Id:SAMEA6504165|INSDC center name:Computational Biology Unit|INSDC first public:2020 03 27T17:05:06Z|INSDC last update:2020 01 27T16:04:35Z|INSDC status:public|Submitter Id:Shield 4Ei|common name:zebrafish|sample name:Shield 4Ei|scientific name:Danio rerio                 NextSeq 500 sequencing ena EXPERIMENT Computational Biology Unit 27 01 2020 16:24:36:406 10 Shield 4Ei OTHER RNA Seq   RNA-Seq TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   ERP116106 NextSeq 500 sequencing ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2020 02 14     1435748376.0 18891426.0 ena RUN Computational Biology Unit 27 01 2020 16:24:36:406 10 0:76 A:489056518;C:372290023;G:393663734;T:180723629;N:14472 76       489056518 372290023 393663734 180723629 14472 ERX3854562 ERS4268611 ERA2359305 Computational Biology Unit|European Nucleotide Archive Computational Biology Unit 1 0.11942   0.03394   0.98971   0.62271   76   B   usable mapping rate illumina nextseq unknown other unknown bulk unknown unknown   Unknown 2019-07-03 Gastrula Embryo Undetermined Embryo Imprecise
9720 9720 ERR3841999 ERX3854561 ERS3556006 ERP116106 PRJEB33323 Deconstructing the individual steps of vertebrate translation initiation ena-STUDY-Computational Biology Unit-03-07-2019-10:11:34:314-422 Other In eukaryotes the number of ribosomes synthesizing a given protein depends on how many are recruited to its mRNA their success in navigating its five prime untranslated region UTR and whether they recognize its start codon. Initiation of translation is a rate limiting step in protein synthesis and key to gene expression control 1 but despite this centrality it remains poorly understood 2 3. Here we introduce ribosome complex profiling RCP seq to capture the transcriptome wide occupancy of scanning initiating elongating and terminating ribosome complexes in a higher eukaryote. We track scanning and elongating ribosomes across all five prime UTRs in zebrafish which enable us to assess the individual regulatory contributions from the three stages of initiation: ribosome recruitment scanning of the five prime UTR and recognition of the start codon. Our data sheds light on small subunit recruitment to mRNAs presenting evidence for the threading model and demonstrates that sequence features regulate this recruitment. We estimate the processivity of scanning ribosomes as they traverse the five prime UTR and show that the repressive effects of upstream open reading frames depend on the efficiency of both translation initiation and termination. Finally we determine the optimal initiation contexts by directly estimating the conversion of scanning to elongating ribosomes and demonstrate specific regulation of translation initiation at the endoplasmic reticulum. Our results open for the possibility of deconvoluting translation initiation into separate stages and provides the first view of global occupancy of ribosomal small subunits in a vertebrate. ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 07 03     Shield 3 SAMEA5752547 Computational Biology Unit ENA FIRST PUBLIC:2020 03 27T17:04:59Z|ENA LAST UPDATE:2019 07 03T10:01:14Z|External Id:SAMEA5752547|INSDC center name:Computational Biology Unit|INSDC first public:2020 03 27T17:04:59Z|INSDC last update:2019 07 03T10:01:14Z|INSDC status:public|Submitter Id:10|common name:zebrafish|dev stage:Shield|sample name:10|scientific name:Danio rerio                 NextSeq 500 sequencing ena EXPERIMENT Computational Biology Unit 27 01 2020 16:24:36:406 9 Shield 3 OTHER RNA Seq   RNA-Seq TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   ERP116106 NextSeq 500 sequencing ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2020 02 14     1168482976.0 15374776.0 ena RUN Computational Biology Unit 27 01 2020 16:24:36:406 9 0:76 A:516070340;C:238363428;G:268806793;T:145231087;N:11328 76       516070340 238363428 268806793 145231087 11328 ERX3854561 ERS3556006 ERA2359305 Computational Biology Unit|European Nucleotide Archive Computational Biology Unit 1 0.22586   0.10275   0.97281   0.47683   76   B   usable mapping rate illumina nextseq unknown other unknown bulk unknown unknown   Unknown 2019-07-03 Gastrula Embryo Undetermined Embryo Imprecise
9721 9721 ERR3841998 ERX3854560 ERS3556007 ERP116106 PRJEB33323 Deconstructing the individual steps of vertebrate translation initiation ena-STUDY-Computational Biology Unit-03-07-2019-10:11:34:314-422 Other In eukaryotes the number of ribosomes synthesizing a given protein depends on how many are recruited to its mRNA their success in navigating its five prime untranslated region UTR and whether they recognize its start codon. Initiation of translation is a rate limiting step in protein synthesis and key to gene expression control 1 but despite this centrality it remains poorly understood 2 3. Here we introduce ribosome complex profiling RCP seq to capture the transcriptome wide occupancy of scanning initiating elongating and terminating ribosome complexes in a higher eukaryote. We track scanning and elongating ribosomes across all five prime UTRs in zebrafish which enable us to assess the individual regulatory contributions from the three stages of initiation: ribosome recruitment scanning of the five prime UTR and recognition of the start codon. Our data sheds light on small subunit recruitment to mRNAs presenting evidence for the threading model and demonstrates that sequence features regulate this recruitment. We estimate the processivity of scanning ribosomes as they traverse the five prime UTR and show that the repressive effects of upstream open reading frames depend on the efficiency of both translation initiation and termination. Finally we determine the optimal initiation contexts by directly estimating the conversion of scanning to elongating ribosomes and demonstrate specific regulation of translation initiation at the endoplasmic reticulum. Our results open for the possibility of deconvoluting translation initiation into separate stages and provides the first view of global occupancy of ribosomal small subunits in a vertebrate. ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 07 03     Shield 4150NT SAMEA5752548 Computational Biology Unit ENA FIRST PUBLIC:2020 03 27T17:04:59Z|ENA LAST UPDATE:2019 07 03T10:01:14Z|External Id:SAMEA5752548|INSDC center name:Computational Biology Unit|INSDC first public:2020 03 27T17:04:59Z|INSDC last update:2019 07 03T10:01:14Z|INSDC status:public|Submitter Id:11|common name:zebrafish|dev stage:Shield|sample name:11|scientific name:Danio rerio                 NextSeq 500 sequencing ena EXPERIMENT Computational Biology Unit 27 01 2020 16:24:36:406 8 Shield 150NT OTHER RNA Seq   RNA-Seq TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   ERP116106 NextSeq 500 sequencing ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2020 02 14     1188343980.0 15636105.0 ena RUN Computational Biology Unit 27 01 2020 16:24:36:406 8 0:76 A:580658556;C:223116826;G:260847322;T:123709681;N:11595 76       580658556 223116826 260847322 123709681 11595 ERX3854560 ERS3556007 ERA2359305 Computational Biology Unit|European Nucleotide Archive Computational Biology Unit 1 0.27037   0.13972   0.97392   0.39459   76   B   usable mapping rate illumina nextseq unknown other unknown bulk unknown unknown   Unknown 2019-07-03 Gastrula Embryo Undetermined Embryo Imprecise
9722 9722 ERR3841997 ERX3854559 ERS3556004 ERP116106 PRJEB33323 Deconstructing the individual steps of vertebrate translation initiation ena-STUDY-Computational Biology Unit-03-07-2019-10:11:34:314-422 Other In eukaryotes the number of ribosomes synthesizing a given protein depends on how many are recruited to its mRNA their success in navigating its five prime untranslated region UTR and whether they recognize its start codon. Initiation of translation is a rate limiting step in protein synthesis and key to gene expression control 1 but despite this centrality it remains poorly understood 2 3. Here we introduce ribosome complex profiling RCP seq to capture the transcriptome wide occupancy of scanning initiating elongating and terminating ribosome complexes in a higher eukaryote. We track scanning and elongating ribosomes across all five prime UTRs in zebrafish which enable us to assess the individual regulatory contributions from the three stages of initiation: ribosome recruitment scanning of the five prime UTR and recognition of the start codon. Our data sheds light on small subunit recruitment to mRNAs presenting evidence for the threading model and demonstrates that sequence features regulate this recruitment. We estimate the processivity of scanning ribosomes as they traverse the five prime UTR and show that the repressive effects of upstream open reading frames depend on the efficiency of both translation initiation and termination. Finally we determine the optimal initiation contexts by directly estimating the conversion of scanning to elongating ribosomes and demonstrate specific regulation of translation initiation at the endoplasmic reticulum. Our results open for the possibility of deconvoluting translation initiation into separate stages and provides the first view of global occupancy of ribosomal small subunits in a vertebrate. ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 07 03     Shield 1 SAMEA5752545 Computational Biology Unit ENA FIRST PUBLIC:2020 03 27T17:04:59Z|ENA LAST UPDATE:2019 07 03T10:01:14Z|External Id:SAMEA5752545|INSDC center name:Computational Biology Unit|INSDC first public:2020 03 27T17:04:59Z|INSDC last update:2019 07 03T10:01:14Z|INSDC status:public|Submitter Id:8|common name:zebrafish|dev stage:Shield|sample name:8|scientific name:Danio rerio                 NextSeq 500 sequencing ena EXPERIMENT Computational Biology Unit 27 01 2020 16:24:36:406 7 Shield 1 OTHER RNA Seq   RNA-Seq TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   ERP116106 NextSeq 500 sequencing ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2020 02 14     1278891444.0 16827519.0 ena RUN Computational Biology Unit 27 01 2020 16:24:36:406 7 0:76 A:571738369;C:256385478;G:295145281;T:155610082;N:12234 76       571738369 256385478 295145281 155610082 12234 ERX3854559 ERS3556004 ERA2359305 Computational Biology Unit|European Nucleotide Archive Computational Biology Unit 1 0.23116   0.11137   0.97932   0.45978   76   B   usable mapping rate illumina nextseq unknown other unknown bulk unknown unknown   Unknown 2019-07-03 Gastrula Embryo Undetermined Embryo Imprecise
9729 9729 ERR3489881 ERX3511296 ERS3556004 ERP116106 PRJEB33323 Deconstructing the individual steps of vertebrate translation initiation ena-STUDY-Computational Biology Unit-03-07-2019-10:11:34:314-422 Other In eukaryotes the number of ribosomes synthesizing a given protein depends on how many are recruited to its mRNA their success in navigating its five prime untranslated region UTR and whether they recognize its start codon. Initiation of translation is a rate limiting step in protein synthesis and key to gene expression control 1 but despite this centrality it remains poorly understood 2 3. Here we introduce ribosome complex profiling RCP seq to capture the transcriptome wide occupancy of scanning initiating elongating and terminating ribosome complexes in a higher eukaryote. We track scanning and elongating ribosomes across all five prime UTRs in zebrafish which enable us to assess the individual regulatory contributions from the three stages of initiation: ribosome recruitment scanning of the five prime UTR and recognition of the start codon. Our data sheds light on small subunit recruitment to mRNAs presenting evidence for the threading model and demonstrates that sequence features regulate this recruitment. We estimate the processivity of scanning ribosomes as they traverse the five prime UTR and show that the repressive effects of upstream open reading frames depend on the efficiency of both translation initiation and termination. Finally we determine the optimal initiation contexts by directly estimating the conversion of scanning to elongating ribosomes and demonstrate specific regulation of translation initiation at the endoplasmic reticulum. Our results open for the possibility of deconvoluting translation initiation into separate stages and provides the first view of global occupancy of ribosomal small subunits in a vertebrate. ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 07 03     Shield 1 SAMEA5752545 Computational Biology Unit ENA FIRST PUBLIC:2020 03 27T17:04:59Z|ENA LAST UPDATE:2019 07 03T10:01:14Z|External Id:SAMEA5752545|INSDC center name:Computational Biology Unit|INSDC first public:2020 03 27T17:04:59Z|INSDC last update:2019 07 03T10:01:14Z|INSDC status:public|Submitter Id:8|common name:zebrafish|dev stage:Shield|sample name:8|scientific name:Danio rerio                 NextSeq 500 sequencing ena EXPERIMENT Computational Biology Unit 22 08 2019 11:57:43:424 33 Shield 1 F20 None RCP seq   OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   ERP116106 NextSeq 500 sequencing ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 08 22     975578636.0 12836561.0 ena RUN Computational Biology Unit 22 08 2019 11:57:43:424 33 0:76 A:398325549;C:237563934;G:230721299;T:108957698;N:10156 76       398325549 237563934 230721299 108957698 10156 ERX3511296 ERS3556004 ERA2100634 Computational Biology Unit|European Nucleotide Archive Computational Biology Unit 1 0.33102   0.19766   0.99918   0.12812   76   B   usable mapping rate illumina nextseq unknown other unknown bulk unknown unknown   Unknown 2019-07-03 Gastrula Embryo Undetermined Embryo Imprecise
9730 9730 ERR3489880 ERX3511295 ERS3556004 ERP116106 PRJEB33323 Deconstructing the individual steps of vertebrate translation initiation ena-STUDY-Computational Biology Unit-03-07-2019-10:11:34:314-422 Other In eukaryotes the number of ribosomes synthesizing a given protein depends on how many are recruited to its mRNA their success in navigating its five prime untranslated region UTR and whether they recognize its start codon. Initiation of translation is a rate limiting step in protein synthesis and key to gene expression control 1 but despite this centrality it remains poorly understood 2 3. Here we introduce ribosome complex profiling RCP seq to capture the transcriptome wide occupancy of scanning initiating elongating and terminating ribosome complexes in a higher eukaryote. We track scanning and elongating ribosomes across all five prime UTRs in zebrafish which enable us to assess the individual regulatory contributions from the three stages of initiation: ribosome recruitment scanning of the five prime UTR and recognition of the start codon. Our data sheds light on small subunit recruitment to mRNAs presenting evidence for the threading model and demonstrates that sequence features regulate this recruitment. We estimate the processivity of scanning ribosomes as they traverse the five prime UTR and show that the repressive effects of upstream open reading frames depend on the efficiency of both translation initiation and termination. Finally we determine the optimal initiation contexts by directly estimating the conversion of scanning to elongating ribosomes and demonstrate specific regulation of translation initiation at the endoplasmic reticulum. Our results open for the possibility of deconvoluting translation initiation into separate stages and provides the first view of global occupancy of ribosomal small subunits in a vertebrate. ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 07 03     Shield 1 SAMEA5752545 Computational Biology Unit ENA FIRST PUBLIC:2020 03 27T17:04:59Z|ENA LAST UPDATE:2019 07 03T10:01:14Z|External Id:SAMEA5752545|INSDC center name:Computational Biology Unit|INSDC first public:2020 03 27T17:04:59Z|INSDC last update:2019 07 03T10:01:14Z|INSDC status:public|Submitter Id:8|common name:zebrafish|dev stage:Shield|sample name:8|scientific name:Danio rerio                 NextSeq 500 sequencing ena EXPERIMENT Computational Biology Unit 22 08 2019 11:57:43:424 32 Shield 1 F19 None RCP seq   OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   ERP116106 NextSeq 500 sequencing ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 08 22     931166668.0 12252193.0 ena RUN Computational Biology Unit 22 08 2019 11:57:43:424 32 0:76 A:271081669;C:253947035;G:272354427;T:133774815;N:8722 76       271081669 253947035 272354427 133774815 8722 ERX3511295 ERS3556004 ERA2100634 Computational Biology Unit|European Nucleotide Archive Computational Biology Unit 1 0.15598   0.10707   0.99902   0.47314   76   B   usable mapping rate illumina nextseq unknown other unknown bulk unknown unknown   Unknown 2019-07-03 Gastrula Embryo Undetermined Embryo Imprecise
9731 9731 ERR3489879 ERX3511294 ERS3556004 ERP116106 PRJEB33323 Deconstructing the individual steps of vertebrate translation initiation ena-STUDY-Computational Biology Unit-03-07-2019-10:11:34:314-422 Other In eukaryotes the number of ribosomes synthesizing a given protein depends on how many are recruited to its mRNA their success in navigating its five prime untranslated region UTR and whether they recognize its start codon. Initiation of translation is a rate limiting step in protein synthesis and key to gene expression control 1 but despite this centrality it remains poorly understood 2 3. Here we introduce ribosome complex profiling RCP seq to capture the transcriptome wide occupancy of scanning initiating elongating and terminating ribosome complexes in a higher eukaryote. We track scanning and elongating ribosomes across all five prime UTRs in zebrafish which enable us to assess the individual regulatory contributions from the three stages of initiation: ribosome recruitment scanning of the five prime UTR and recognition of the start codon. Our data sheds light on small subunit recruitment to mRNAs presenting evidence for the threading model and demonstrates that sequence features regulate this recruitment. We estimate the processivity of scanning ribosomes as they traverse the five prime UTR and show that the repressive effects of upstream open reading frames depend on the efficiency of both translation initiation and termination. Finally we determine the optimal initiation contexts by directly estimating the conversion of scanning to elongating ribosomes and demonstrate specific regulation of translation initiation at the endoplasmic reticulum. Our results open for the possibility of deconvoluting translation initiation into separate stages and provides the first view of global occupancy of ribosomal small subunits in a vertebrate. ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 07 03     Shield 1 SAMEA5752545 Computational Biology Unit ENA FIRST PUBLIC:2020 03 27T17:04:59Z|ENA LAST UPDATE:2019 07 03T10:01:14Z|External Id:SAMEA5752545|INSDC center name:Computational Biology Unit|INSDC first public:2020 03 27T17:04:59Z|INSDC last update:2019 07 03T10:01:14Z|INSDC status:public|Submitter Id:8|common name:zebrafish|dev stage:Shield|sample name:8|scientific name:Danio rerio                 NextSeq 500 sequencing ena EXPERIMENT Computational Biology Unit 22 08 2019 11:57:43:424 31 Shield 1 F18 None RCP seq   OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   ERP116106 NextSeq 500 sequencing ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 08 22     1506513268.0 19822543.0 ena RUN Computational Biology Unit 22 08 2019 11:57:43:424 31 0:76 A:493273515;C:456544968;G:391677033;T:165002559;N:15193 76       493273515 456544968 391677033 165002559 15193 ERX3511294 ERS3556004 ERA2100634 Computational Biology Unit|European Nucleotide Archive Computational Biology Unit 1 0.01562   0.0053   0.99908   0.8127   76   B   usable mapping rate illumina nextseq unknown other unknown bulk unknown unknown   Unknown 2019-07-03 Gastrula Embryo Undetermined Embryo Imprecise
9732 9732 ERR3489878 ERX3511293 ERS3556004 ERP116106 PRJEB33323 Deconstructing the individual steps of vertebrate translation initiation ena-STUDY-Computational Biology Unit-03-07-2019-10:11:34:314-422 Other In eukaryotes the number of ribosomes synthesizing a given protein depends on how many are recruited to its mRNA their success in navigating its five prime untranslated region UTR and whether they recognize its start codon. Initiation of translation is a rate limiting step in protein synthesis and key to gene expression control 1 but despite this centrality it remains poorly understood 2 3. Here we introduce ribosome complex profiling RCP seq to capture the transcriptome wide occupancy of scanning initiating elongating and terminating ribosome complexes in a higher eukaryote. We track scanning and elongating ribosomes across all five prime UTRs in zebrafish which enable us to assess the individual regulatory contributions from the three stages of initiation: ribosome recruitment scanning of the five prime UTR and recognition of the start codon. Our data sheds light on small subunit recruitment to mRNAs presenting evidence for the threading model and demonstrates that sequence features regulate this recruitment. We estimate the processivity of scanning ribosomes as they traverse the five prime UTR and show that the repressive effects of upstream open reading frames depend on the efficiency of both translation initiation and termination. Finally we determine the optimal initiation contexts by directly estimating the conversion of scanning to elongating ribosomes and demonstrate specific regulation of translation initiation at the endoplasmic reticulum. Our results open for the possibility of deconvoluting translation initiation into separate stages and provides the first view of global occupancy of ribosomal small subunits in a vertebrate. ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 07 03     Shield 1 SAMEA5752545 Computational Biology Unit ENA FIRST PUBLIC:2020 03 27T17:04:59Z|ENA LAST UPDATE:2019 07 03T10:01:14Z|External Id:SAMEA5752545|INSDC center name:Computational Biology Unit|INSDC first public:2020 03 27T17:04:59Z|INSDC last update:2019 07 03T10:01:14Z|INSDC status:public|Submitter Id:8|common name:zebrafish|dev stage:Shield|sample name:8|scientific name:Danio rerio                 NextSeq 500 sequencing ena EXPERIMENT Computational Biology Unit 22 08 2019 11:57:43:424 30 Shield 1 F17 None RCP seq   OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   ERP116106 NextSeq 500 sequencing ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 08 22     1259456496.0 16571796.0 ena RUN Computational Biology Unit 22 08 2019 11:57:43:424 30 0:76 A:471473123;C:333726472;G:302016190;T:152228002;N:12709 76       471473123 333726472 302016190 152228002 12709 ERX3511293 ERS3556004 ERA2100634 Computational Biology Unit|European Nucleotide Archive Computational Biology Unit 1 0.18339   0.12544   0.99928   0.22368   76   B   usable mapping rate illumina nextseq unknown other unknown bulk unknown unknown   Unknown 2019-07-03 Gastrula Embryo Undetermined Embryo Imprecise
9733 9733 ERR3489877 ERX3511292 ERS3556004 ERP116106 PRJEB33323 Deconstructing the individual steps of vertebrate translation initiation ena-STUDY-Computational Biology Unit-03-07-2019-10:11:34:314-422 Other In eukaryotes the number of ribosomes synthesizing a given protein depends on how many are recruited to its mRNA their success in navigating its five prime untranslated region UTR and whether they recognize its start codon. Initiation of translation is a rate limiting step in protein synthesis and key to gene expression control 1 but despite this centrality it remains poorly understood 2 3. Here we introduce ribosome complex profiling RCP seq to capture the transcriptome wide occupancy of scanning initiating elongating and terminating ribosome complexes in a higher eukaryote. We track scanning and elongating ribosomes across all five prime UTRs in zebrafish which enable us to assess the individual regulatory contributions from the three stages of initiation: ribosome recruitment scanning of the five prime UTR and recognition of the start codon. Our data sheds light on small subunit recruitment to mRNAs presenting evidence for the threading model and demonstrates that sequence features regulate this recruitment. We estimate the processivity of scanning ribosomes as they traverse the five prime UTR and show that the repressive effects of upstream open reading frames depend on the efficiency of both translation initiation and termination. Finally we determine the optimal initiation contexts by directly estimating the conversion of scanning to elongating ribosomes and demonstrate specific regulation of translation initiation at the endoplasmic reticulum. Our results open for the possibility of deconvoluting translation initiation into separate stages and provides the first view of global occupancy of ribosomal small subunits in a vertebrate. ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 07 03     Shield 1 SAMEA5752545 Computational Biology Unit ENA FIRST PUBLIC:2020 03 27T17:04:59Z|ENA LAST UPDATE:2019 07 03T10:01:14Z|External Id:SAMEA5752545|INSDC center name:Computational Biology Unit|INSDC first public:2020 03 27T17:04:59Z|INSDC last update:2019 07 03T10:01:14Z|INSDC status:public|Submitter Id:8|common name:zebrafish|dev stage:Shield|sample name:8|scientific name:Danio rerio                 NextSeq 500 sequencing ena EXPERIMENT Computational Biology Unit 22 08 2019 11:57:43:424 29 Shield 1 F16 None RCP seq   OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   ERP116106 NextSeq 500 sequencing ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 08 22     1364615872.0 17955472.0 ena RUN Computational Biology Unit 22 08 2019 11:57:43:424 29 0:76 A:539462776;C:341141880;G:314571683;T:169426048;N:13485 76       539462776 341141880 314571683 169426048 13485 ERX3511292 ERS3556004 ERA2100634 Computational Biology Unit|European Nucleotide Archive Computational Biology Unit 1 0.11663   0.07319   0.99939   0.25377   76   B   usable mapping rate illumina nextseq unknown other unknown bulk unknown unknown   Unknown 2019-07-03 Gastrula Embryo Undetermined Embryo Imprecise
9734 9734 ERR3489876 ERX3511291 ERS3556004 ERP116106 PRJEB33323 Deconstructing the individual steps of vertebrate translation initiation ena-STUDY-Computational Biology Unit-03-07-2019-10:11:34:314-422 Other In eukaryotes the number of ribosomes synthesizing a given protein depends on how many are recruited to its mRNA their success in navigating its five prime untranslated region UTR and whether they recognize its start codon. Initiation of translation is a rate limiting step in protein synthesis and key to gene expression control 1 but despite this centrality it remains poorly understood 2 3. Here we introduce ribosome complex profiling RCP seq to capture the transcriptome wide occupancy of scanning initiating elongating and terminating ribosome complexes in a higher eukaryote. We track scanning and elongating ribosomes across all five prime UTRs in zebrafish which enable us to assess the individual regulatory contributions from the three stages of initiation: ribosome recruitment scanning of the five prime UTR and recognition of the start codon. Our data sheds light on small subunit recruitment to mRNAs presenting evidence for the threading model and demonstrates that sequence features regulate this recruitment. We estimate the processivity of scanning ribosomes as they traverse the five prime UTR and show that the repressive effects of upstream open reading frames depend on the efficiency of both translation initiation and termination. Finally we determine the optimal initiation contexts by directly estimating the conversion of scanning to elongating ribosomes and demonstrate specific regulation of translation initiation at the endoplasmic reticulum. Our results open for the possibility of deconvoluting translation initiation into separate stages and provides the first view of global occupancy of ribosomal small subunits in a vertebrate. ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 07 03     Shield 1 SAMEA5752545 Computational Biology Unit ENA FIRST PUBLIC:2020 03 27T17:04:59Z|ENA LAST UPDATE:2019 07 03T10:01:14Z|External Id:SAMEA5752545|INSDC center name:Computational Biology Unit|INSDC first public:2020 03 27T17:04:59Z|INSDC last update:2019 07 03T10:01:14Z|INSDC status:public|Submitter Id:8|common name:zebrafish|dev stage:Shield|sample name:8|scientific name:Danio rerio                 NextSeq 500 sequencing ena EXPERIMENT Computational Biology Unit 22 08 2019 11:57:43:424 28 Shield 1 F15 None RCP seq   OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   ERP116106 NextSeq 500 sequencing ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 08 22     952605888.0 12534288.0 ena RUN Computational Biology Unit 22 08 2019 11:57:43:424 28 0:76 A:414133320;C:219437277;G:207418049;T:111607418;N:9824 76       414133320 219437277 207418049 111607418 9824 ERX3511291 ERS3556004 ERA2100634 Computational Biology Unit|European Nucleotide Archive Computational Biology Unit 1 0.17603   0.10972   0.99935   0.13311   76   B   usable mapping rate illumina nextseq unknown other unknown bulk unknown unknown   Unknown 2019-07-03 Gastrula Embryo Undetermined Embryo Imprecise
9735 9735 ERR3489875 ERX3511290 ERS3556004 ERP116106 PRJEB33323 Deconstructing the individual steps of vertebrate translation initiation ena-STUDY-Computational Biology Unit-03-07-2019-10:11:34:314-422 Other In eukaryotes the number of ribosomes synthesizing a given protein depends on how many are recruited to its mRNA their success in navigating its five prime untranslated region UTR and whether they recognize its start codon. Initiation of translation is a rate limiting step in protein synthesis and key to gene expression control 1 but despite this centrality it remains poorly understood 2 3. Here we introduce ribosome complex profiling RCP seq to capture the transcriptome wide occupancy of scanning initiating elongating and terminating ribosome complexes in a higher eukaryote. We track scanning and elongating ribosomes across all five prime UTRs in zebrafish which enable us to assess the individual regulatory contributions from the three stages of initiation: ribosome recruitment scanning of the five prime UTR and recognition of the start codon. Our data sheds light on small subunit recruitment to mRNAs presenting evidence for the threading model and demonstrates that sequence features regulate this recruitment. We estimate the processivity of scanning ribosomes as they traverse the five prime UTR and show that the repressive effects of upstream open reading frames depend on the efficiency of both translation initiation and termination. Finally we determine the optimal initiation contexts by directly estimating the conversion of scanning to elongating ribosomes and demonstrate specific regulation of translation initiation at the endoplasmic reticulum. Our results open for the possibility of deconvoluting translation initiation into separate stages and provides the first view of global occupancy of ribosomal small subunits in a vertebrate. ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 07 03     Shield 1 SAMEA5752545 Computational Biology Unit ENA FIRST PUBLIC:2020 03 27T17:04:59Z|ENA LAST UPDATE:2019 07 03T10:01:14Z|External Id:SAMEA5752545|INSDC center name:Computational Biology Unit|INSDC first public:2020 03 27T17:04:59Z|INSDC last update:2019 07 03T10:01:14Z|INSDC status:public|Submitter Id:8|common name:zebrafish|dev stage:Shield|sample name:8|scientific name:Danio rerio                 NextSeq 500 sequencing ena EXPERIMENT Computational Biology Unit 22 08 2019 11:57:43:424 27 Shield 1 F14 None RCP seq   OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   ERP116106 NextSeq 500 sequencing ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 08 22     870952628.0 11459903.0 ena RUN Computational Biology Unit 22 08 2019 11:57:43:424 27 0:76 A:357710338;C:221475453;G:191231014;T:100526675;N:9148 76       357710338 221475453 191231014 100526675 9148 ERX3511290 ERS3556004 ERA2100634 Computational Biology Unit|European Nucleotide Archive Computational Biology Unit 1 0.1248   0.06422   0.99896   0.34819   76   B   usable mapping rate illumina nextseq unknown other unknown bulk unknown unknown   Unknown 2019-07-03 Gastrula Embryo Undetermined Embryo Imprecise
9736 9736 ERR3489874 ERX3511289 ERS3556004 ERP116106 PRJEB33323 Deconstructing the individual steps of vertebrate translation initiation ena-STUDY-Computational Biology Unit-03-07-2019-10:11:34:314-422 Other In eukaryotes the number of ribosomes synthesizing a given protein depends on how many are recruited to its mRNA their success in navigating its five prime untranslated region UTR and whether they recognize its start codon. Initiation of translation is a rate limiting step in protein synthesis and key to gene expression control 1 but despite this centrality it remains poorly understood 2 3. Here we introduce ribosome complex profiling RCP seq to capture the transcriptome wide occupancy of scanning initiating elongating and terminating ribosome complexes in a higher eukaryote. We track scanning and elongating ribosomes across all five prime UTRs in zebrafish which enable us to assess the individual regulatory contributions from the three stages of initiation: ribosome recruitment scanning of the five prime UTR and recognition of the start codon. Our data sheds light on small subunit recruitment to mRNAs presenting evidence for the threading model and demonstrates that sequence features regulate this recruitment. We estimate the processivity of scanning ribosomes as they traverse the five prime UTR and show that the repressive effects of upstream open reading frames depend on the efficiency of both translation initiation and termination. Finally we determine the optimal initiation contexts by directly estimating the conversion of scanning to elongating ribosomes and demonstrate specific regulation of translation initiation at the endoplasmic reticulum. Our results open for the possibility of deconvoluting translation initiation into separate stages and provides the first view of global occupancy of ribosomal small subunits in a vertebrate. ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 07 03     Shield 1 SAMEA5752545 Computational Biology Unit ENA FIRST PUBLIC:2020 03 27T17:04:59Z|ENA LAST UPDATE:2019 07 03T10:01:14Z|External Id:SAMEA5752545|INSDC center name:Computational Biology Unit|INSDC first public:2020 03 27T17:04:59Z|INSDC last update:2019 07 03T10:01:14Z|INSDC status:public|Submitter Id:8|common name:zebrafish|dev stage:Shield|sample name:8|scientific name:Danio rerio                 NextSeq 500 sequencing ena EXPERIMENT Computational Biology Unit 22 08 2019 11:57:43:424 26 Shield 1 F13 None RCP seq   OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   ERP116106 NextSeq 500 sequencing ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 08 22     981672620.0 12916745.0 ena RUN Computational Biology Unit 22 08 2019 11:57:43:424 26 0:76 A:434153198;C:223317075;G:198260771;T:125932145;N:9431 76       434153198 223317075 198260771 125932145 9431 ERX3511289 ERS3556004 ERA2100634 Computational Biology Unit|European Nucleotide Archive Computational Biology Unit 1 0.44603   0.25602   0.99874   0.18074   76   B   usable mapping rate illumina nextseq unknown other unknown bulk unknown unknown   Unknown 2019-07-03 Gastrula Embryo Undetermined Embryo Imprecise
9737 9737 ERR3489873 ERX3511288 ERS3556004 ERP116106 PRJEB33323 Deconstructing the individual steps of vertebrate translation initiation ena-STUDY-Computational Biology Unit-03-07-2019-10:11:34:314-422 Other In eukaryotes the number of ribosomes synthesizing a given protein depends on how many are recruited to its mRNA their success in navigating its five prime untranslated region UTR and whether they recognize its start codon. Initiation of translation is a rate limiting step in protein synthesis and key to gene expression control 1 but despite this centrality it remains poorly understood 2 3. Here we introduce ribosome complex profiling RCP seq to capture the transcriptome wide occupancy of scanning initiating elongating and terminating ribosome complexes in a higher eukaryote. We track scanning and elongating ribosomes across all five prime UTRs in zebrafish which enable us to assess the individual regulatory contributions from the three stages of initiation: ribosome recruitment scanning of the five prime UTR and recognition of the start codon. Our data sheds light on small subunit recruitment to mRNAs presenting evidence for the threading model and demonstrates that sequence features regulate this recruitment. We estimate the processivity of scanning ribosomes as they traverse the five prime UTR and show that the repressive effects of upstream open reading frames depend on the efficiency of both translation initiation and termination. Finally we determine the optimal initiation contexts by directly estimating the conversion of scanning to elongating ribosomes and demonstrate specific regulation of translation initiation at the endoplasmic reticulum. Our results open for the possibility of deconvoluting translation initiation into separate stages and provides the first view of global occupancy of ribosomal small subunits in a vertebrate. ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 07 03     Shield 1 SAMEA5752545 Computational Biology Unit ENA FIRST PUBLIC:2020 03 27T17:04:59Z|ENA LAST UPDATE:2019 07 03T10:01:14Z|External Id:SAMEA5752545|INSDC center name:Computational Biology Unit|INSDC first public:2020 03 27T17:04:59Z|INSDC last update:2019 07 03T10:01:14Z|INSDC status:public|Submitter Id:8|common name:zebrafish|dev stage:Shield|sample name:8|scientific name:Danio rerio                 NextSeq 500 sequencing ena EXPERIMENT Computational Biology Unit 22 08 2019 11:57:43:424 25 Shield 1 F12 None RCP seq   OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   ERP116106 NextSeq 500 sequencing ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 08 22     1304618128.0 17166028.0 ena RUN Computational Biology Unit 22 08 2019 11:57:43:424 25 0:76 A:651341516;C:270458496;G:243929520;T:138874830;N:13766 76       651341516 270458496 243929520 138874830 13766 ERX3511288 ERS3556004 ERA2100634 Computational Biology Unit|European Nucleotide Archive Computational Biology Unit 1 0.64293   0.38159   0.99886   0.07313   76   B   usable mapping rate illumina nextseq unknown other unknown bulk unknown unknown   Unknown 2019-07-03 Gastrula Embryo Undetermined Embryo Imprecise
9738 9738 ERR3489872 ERX3511287 ERS3556004 ERP116106 PRJEB33323 Deconstructing the individual steps of vertebrate translation initiation ena-STUDY-Computational Biology Unit-03-07-2019-10:11:34:314-422 Other In eukaryotes the number of ribosomes synthesizing a given protein depends on how many are recruited to its mRNA their success in navigating its five prime untranslated region UTR and whether they recognize its start codon. Initiation of translation is a rate limiting step in protein synthesis and key to gene expression control 1 but despite this centrality it remains poorly understood 2 3. Here we introduce ribosome complex profiling RCP seq to capture the transcriptome wide occupancy of scanning initiating elongating and terminating ribosome complexes in a higher eukaryote. We track scanning and elongating ribosomes across all five prime UTRs in zebrafish which enable us to assess the individual regulatory contributions from the three stages of initiation: ribosome recruitment scanning of the five prime UTR and recognition of the start codon. Our data sheds light on small subunit recruitment to mRNAs presenting evidence for the threading model and demonstrates that sequence features regulate this recruitment. We estimate the processivity of scanning ribosomes as they traverse the five prime UTR and show that the repressive effects of upstream open reading frames depend on the efficiency of both translation initiation and termination. Finally we determine the optimal initiation contexts by directly estimating the conversion of scanning to elongating ribosomes and demonstrate specific regulation of translation initiation at the endoplasmic reticulum. Our results open for the possibility of deconvoluting translation initiation into separate stages and provides the first view of global occupancy of ribosomal small subunits in a vertebrate. ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 07 03     Shield 1 SAMEA5752545 Computational Biology Unit ENA FIRST PUBLIC:2020 03 27T17:04:59Z|ENA LAST UPDATE:2019 07 03T10:01:14Z|External Id:SAMEA5752545|INSDC center name:Computational Biology Unit|INSDC first public:2020 03 27T17:04:59Z|INSDC last update:2019 07 03T10:01:14Z|INSDC status:public|Submitter Id:8|common name:zebrafish|dev stage:Shield|sample name:8|scientific name:Danio rerio                 NextSeq 500 sequencing ena EXPERIMENT Computational Biology Unit 22 08 2019 11:57:43:424 24 Shield 1 F10 None RCP seq   OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   ERP116106 NextSeq 500 sequencing ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 08 22     1336115948.0 17580473.0 ena RUN Computational Biology Unit 22 08 2019 11:57:43:424 24 0:76 A:608634206;C:295943144;G:286494431;T:145029697;N:14470 76       608634206 295943144 286494431 145029697 14470 ERX3511287 ERS3556004 ERA2100634 Computational Biology Unit|European Nucleotide Archive Computational Biology Unit 1 0.68758   0.47517   0.99898   0.02301   76   B   usable mapping rate illumina nextseq unknown other unknown bulk unknown unknown   Unknown 2019-07-03 Gastrula Embryo Undetermined Embryo Imprecise
9739 9739 ERR3489871 ERX3511286 ERS3556004 ERP116106 PRJEB33323 Deconstructing the individual steps of vertebrate translation initiation ena-STUDY-Computational Biology Unit-03-07-2019-10:11:34:314-422 Other In eukaryotes the number of ribosomes synthesizing a given protein depends on how many are recruited to its mRNA their success in navigating its five prime untranslated region UTR and whether they recognize its start codon. Initiation of translation is a rate limiting step in protein synthesis and key to gene expression control 1 but despite this centrality it remains poorly understood 2 3. Here we introduce ribosome complex profiling RCP seq to capture the transcriptome wide occupancy of scanning initiating elongating and terminating ribosome complexes in a higher eukaryote. We track scanning and elongating ribosomes across all five prime UTRs in zebrafish which enable us to assess the individual regulatory contributions from the three stages of initiation: ribosome recruitment scanning of the five prime UTR and recognition of the start codon. Our data sheds light on small subunit recruitment to mRNAs presenting evidence for the threading model and demonstrates that sequence features regulate this recruitment. We estimate the processivity of scanning ribosomes as they traverse the five prime UTR and show that the repressive effects of upstream open reading frames depend on the efficiency of both translation initiation and termination. Finally we determine the optimal initiation contexts by directly estimating the conversion of scanning to elongating ribosomes and demonstrate specific regulation of translation initiation at the endoplasmic reticulum. Our results open for the possibility of deconvoluting translation initiation into separate stages and provides the first view of global occupancy of ribosomal small subunits in a vertebrate. ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 07 03     Shield 1 SAMEA5752545 Computational Biology Unit ENA FIRST PUBLIC:2020 03 27T17:04:59Z|ENA LAST UPDATE:2019 07 03T10:01:14Z|External Id:SAMEA5752545|INSDC center name:Computational Biology Unit|INSDC first public:2020 03 27T17:04:59Z|INSDC last update:2019 07 03T10:01:14Z|INSDC status:public|Submitter Id:8|common name:zebrafish|dev stage:Shield|sample name:8|scientific name:Danio rerio                 NextSeq 500 sequencing ena EXPERIMENT Computational Biology Unit 22 08 2019 11:57:43:424 23 Shield 1 F9 None RCP seq   OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   ERP116106 NextSeq 500 sequencing ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 08 22     1434402492.0 18873717.0 ena RUN Computational Biology Unit 22 08 2019 11:57:43:424 23 0:76 A:658705664;C:297967081;G:295595736;T:182118632;N:15379 76       658705664 297967081 295595736 182118632 15379 ERX3511286 ERS3556004 ERA2100634 Computational Biology Unit|European Nucleotide Archive Computational Biology Unit 1 0.74246   0.44235   0.99701   0.03112   76   B   usable mapping rate illumina nextseq unknown other unknown bulk unknown unknown   Unknown 2019-07-03 Gastrula Embryo Undetermined Embryo Imprecise
9740 9740 ERR3489870 ERX3511285 ERS3556007 ERP116106 PRJEB33323 Deconstructing the individual steps of vertebrate translation initiation ena-STUDY-Computational Biology Unit-03-07-2019-10:11:34:314-422 Other In eukaryotes the number of ribosomes synthesizing a given protein depends on how many are recruited to its mRNA their success in navigating its five prime untranslated region UTR and whether they recognize its start codon. Initiation of translation is a rate limiting step in protein synthesis and key to gene expression control 1 but despite this centrality it remains poorly understood 2 3. Here we introduce ribosome complex profiling RCP seq to capture the transcriptome wide occupancy of scanning initiating elongating and terminating ribosome complexes in a higher eukaryote. We track scanning and elongating ribosomes across all five prime UTRs in zebrafish which enable us to assess the individual regulatory contributions from the three stages of initiation: ribosome recruitment scanning of the five prime UTR and recognition of the start codon. Our data sheds light on small subunit recruitment to mRNAs presenting evidence for the threading model and demonstrates that sequence features regulate this recruitment. We estimate the processivity of scanning ribosomes as they traverse the five prime UTR and show that the repressive effects of upstream open reading frames depend on the efficiency of both translation initiation and termination. Finally we determine the optimal initiation contexts by directly estimating the conversion of scanning to elongating ribosomes and demonstrate specific regulation of translation initiation at the endoplasmic reticulum. Our results open for the possibility of deconvoluting translation initiation into separate stages and provides the first view of global occupancy of ribosomal small subunits in a vertebrate. ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 07 03     Shield 4150NT SAMEA5752548 Computational Biology Unit ENA FIRST PUBLIC:2020 03 27T17:04:59Z|ENA LAST UPDATE:2019 07 03T10:01:14Z|External Id:SAMEA5752548|INSDC center name:Computational Biology Unit|INSDC first public:2020 03 27T17:04:59Z|INSDC last update:2019 07 03T10:01:14Z|INSDC status:public|Submitter Id:11|common name:zebrafish|dev stage:Shield|sample name:11|scientific name:Danio rerio                 NextSeq 500 sequencing ena EXPERIMENT Computational Biology Unit 22 08 2019 11:57:43:424 22 Shield 4150NT LSU None RCP seq   OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   ERP116106 NextSeq 500 sequencing ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 08 24     24063208094.0 159358994.0 ena RUN Computational Biology Unit 22 08 2019 11:57:43:424 22 0:151 A:7153064588;C:5242791119;G:8513736630;T:3152547276;N:1068481 151       7153064588 5242791119 8513736630 3152547276 1068481 ERX3511285 ERS3556007 ERA2100634 Computational Biology Unit|European Nucleotide Archive Computational Biology Unit 1 0.81267   0.27138   0.99868   0.91938   151   B   usable mapping rate illumina nextseq unknown other unknown bulk unknown unknown   Unknown 2019-07-03 Gastrula Embryo Undetermined Embryo Imprecise
9741 9741 ERR3489869 ERX3511284 ERS3556007 ERP116106 PRJEB33323 Deconstructing the individual steps of vertebrate translation initiation ena-STUDY-Computational Biology Unit-03-07-2019-10:11:34:314-422 Other In eukaryotes the number of ribosomes synthesizing a given protein depends on how many are recruited to its mRNA their success in navigating its five prime untranslated region UTR and whether they recognize its start codon. Initiation of translation is a rate limiting step in protein synthesis and key to gene expression control 1 but despite this centrality it remains poorly understood 2 3. Here we introduce ribosome complex profiling RCP seq to capture the transcriptome wide occupancy of scanning initiating elongating and terminating ribosome complexes in a higher eukaryote. We track scanning and elongating ribosomes across all five prime UTRs in zebrafish which enable us to assess the individual regulatory contributions from the three stages of initiation: ribosome recruitment scanning of the five prime UTR and recognition of the start codon. Our data sheds light on small subunit recruitment to mRNAs presenting evidence for the threading model and demonstrates that sequence features regulate this recruitment. We estimate the processivity of scanning ribosomes as they traverse the five prime UTR and show that the repressive effects of upstream open reading frames depend on the efficiency of both translation initiation and termination. Finally we determine the optimal initiation contexts by directly estimating the conversion of scanning to elongating ribosomes and demonstrate specific regulation of translation initiation at the endoplasmic reticulum. Our results open for the possibility of deconvoluting translation initiation into separate stages and provides the first view of global occupancy of ribosomal small subunits in a vertebrate. ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 07 03     Shield 4150NT SAMEA5752548 Computational Biology Unit ENA FIRST PUBLIC:2020 03 27T17:04:59Z|ENA LAST UPDATE:2019 07 03T10:01:14Z|External Id:SAMEA5752548|INSDC center name:Computational Biology Unit|INSDC first public:2020 03 27T17:04:59Z|INSDC last update:2019 07 03T10:01:14Z|INSDC status:public|Submitter Id:11|common name:zebrafish|dev stage:Shield|sample name:11|scientific name:Danio rerio                 NextSeq 500 sequencing ena EXPERIMENT Computational Biology Unit 22 08 2019 11:57:43:424 21 Shield 4150NT SSU None RCP seq   OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   ERP116106 NextSeq 500 sequencing ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 08 24     14987998619.0 99258269.0 ena RUN Computational Biology Unit 22 08 2019 11:57:43:424 21 0:151 A:4578051806;C:2616328922;G:5914185813;T:1878780090;N:651988 151       4578051806 2616328922 5914185813 1878780090 651988 ERX3511284 ERS3556007 ERA2100634 Computational Biology Unit|European Nucleotide Archive Computational Biology Unit 1 0.77096   0.5278   0.99833   0.42635   151   B   usable mapping rate illumina nextseq unknown other unknown bulk unknown unknown   Unknown 2019-07-03 Gastrula Embryo Undetermined Embryo Imprecise
9746 9746 ERR3489864 ERX3511279 ERS3556006 ERP116106 PRJEB33323 Deconstructing the individual steps of vertebrate translation initiation ena-STUDY-Computational Biology Unit-03-07-2019-10:11:34:314-422 Other In eukaryotes the number of ribosomes synthesizing a given protein depends on how many are recruited to its mRNA their success in navigating its five prime untranslated region UTR and whether they recognize its start codon. Initiation of translation is a rate limiting step in protein synthesis and key to gene expression control 1 but despite this centrality it remains poorly understood 2 3. Here we introduce ribosome complex profiling RCP seq to capture the transcriptome wide occupancy of scanning initiating elongating and terminating ribosome complexes in a higher eukaryote. We track scanning and elongating ribosomes across all five prime UTRs in zebrafish which enable us to assess the individual regulatory contributions from the three stages of initiation: ribosome recruitment scanning of the five prime UTR and recognition of the start codon. Our data sheds light on small subunit recruitment to mRNAs presenting evidence for the threading model and demonstrates that sequence features regulate this recruitment. We estimate the processivity of scanning ribosomes as they traverse the five prime UTR and show that the repressive effects of upstream open reading frames depend on the efficiency of both translation initiation and termination. Finally we determine the optimal initiation contexts by directly estimating the conversion of scanning to elongating ribosomes and demonstrate specific regulation of translation initiation at the endoplasmic reticulum. Our results open for the possibility of deconvoluting translation initiation into separate stages and provides the first view of global occupancy of ribosomal small subunits in a vertebrate. ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 07 03     Shield 3 SAMEA5752547 Computational Biology Unit ENA FIRST PUBLIC:2020 03 27T17:04:59Z|ENA LAST UPDATE:2019 07 03T10:01:14Z|External Id:SAMEA5752547|INSDC center name:Computational Biology Unit|INSDC first public:2020 03 27T17:04:59Z|INSDC last update:2019 07 03T10:01:14Z|INSDC status:public|Submitter Id:10|common name:zebrafish|dev stage:Shield|sample name:10|scientific name:Danio rerio                 NextSeq 500 sequencing ena EXPERIMENT Computational Biology Unit 22 08 2019 11:57:43:424 16 Shield 3 LSU None RCP seq   OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   ERP116106 NextSeq 500 sequencing ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 08 22     11777220072.0 154963422.0 ena RUN Computational Biology Unit 22 08 2019 11:57:43:424 16 0:76 A:3368038444;C:3190496935;G:3529701152;T:1688766119;N:217422 76       3368038444 3190496935 3529701152 1688766119 217422 ERX3511279 ERS3556006 ERA2100634 Computational Biology Unit|European Nucleotide Archive Computational Biology Unit 1 0.70681   0.18846   0.99332   0.71978   76   B   usable mapping rate illumina nextseq unknown other unknown bulk unknown unknown   Unknown 2019-07-03 Gastrula Embryo Undetermined Embryo Imprecise
9747 9747 ERR3489863 ERX3511278 ERS3556006 ERP116106 PRJEB33323 Deconstructing the individual steps of vertebrate translation initiation ena-STUDY-Computational Biology Unit-03-07-2019-10:11:34:314-422 Other In eukaryotes the number of ribosomes synthesizing a given protein depends on how many are recruited to its mRNA their success in navigating its five prime untranslated region UTR and whether they recognize its start codon. Initiation of translation is a rate limiting step in protein synthesis and key to gene expression control 1 but despite this centrality it remains poorly understood 2 3. Here we introduce ribosome complex profiling RCP seq to capture the transcriptome wide occupancy of scanning initiating elongating and terminating ribosome complexes in a higher eukaryote. We track scanning and elongating ribosomes across all five prime UTRs in zebrafish which enable us to assess the individual regulatory contributions from the three stages of initiation: ribosome recruitment scanning of the five prime UTR and recognition of the start codon. Our data sheds light on small subunit recruitment to mRNAs presenting evidence for the threading model and demonstrates that sequence features regulate this recruitment. We estimate the processivity of scanning ribosomes as they traverse the five prime UTR and show that the repressive effects of upstream open reading frames depend on the efficiency of both translation initiation and termination. Finally we determine the optimal initiation contexts by directly estimating the conversion of scanning to elongating ribosomes and demonstrate specific regulation of translation initiation at the endoplasmic reticulum. Our results open for the possibility of deconvoluting translation initiation into separate stages and provides the first view of global occupancy of ribosomal small subunits in a vertebrate. ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 07 03     Shield 3 SAMEA5752547 Computational Biology Unit ENA FIRST PUBLIC:2020 03 27T17:04:59Z|ENA LAST UPDATE:2019 07 03T10:01:14Z|External Id:SAMEA5752547|INSDC center name:Computational Biology Unit|INSDC first public:2020 03 27T17:04:59Z|INSDC last update:2019 07 03T10:01:14Z|INSDC status:public|Submitter Id:10|common name:zebrafish|dev stage:Shield|sample name:10|scientific name:Danio rerio                 NextSeq 500 sequencing ena EXPERIMENT Computational Biology Unit 22 08 2019 11:57:43:424 15 Shield 3 SSU None RCP seq   OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   ERP116106 NextSeq 500 sequencing ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 08 22     7920419952.0 104216052.0 ena RUN Computational Biology Unit 22 08 2019 11:57:43:424 15 0:76 A:2990167185;C:1767708808;G:2104830363;T:1057568560;N:145036 76       2990167185 1767708808 2104830363 1057568560 145036 ERX3511278 ERS3556006 ERA2100634 Computational Biology Unit|European Nucleotide Archive Computational Biology Unit 1 0.5052   0.30841   0.99129   0.60948   76   B   usable mapping rate illumina nextseq unknown other unknown bulk unknown unknown   Unknown 2019-07-03 Gastrula Embryo Undetermined Embryo Imprecise
9748 9748 ERR3489862 ERX3511277 ERS3556005 ERP116106 PRJEB33323 Deconstructing the individual steps of vertebrate translation initiation ena-STUDY-Computational Biology Unit-03-07-2019-10:11:34:314-422 Other In eukaryotes the number of ribosomes synthesizing a given protein depends on how many are recruited to its mRNA their success in navigating its five prime untranslated region UTR and whether they recognize its start codon. Initiation of translation is a rate limiting step in protein synthesis and key to gene expression control 1 but despite this centrality it remains poorly understood 2 3. Here we introduce ribosome complex profiling RCP seq to capture the transcriptome wide occupancy of scanning initiating elongating and terminating ribosome complexes in a higher eukaryote. We track scanning and elongating ribosomes across all five prime UTRs in zebrafish which enable us to assess the individual regulatory contributions from the three stages of initiation: ribosome recruitment scanning of the five prime UTR and recognition of the start codon. Our data sheds light on small subunit recruitment to mRNAs presenting evidence for the threading model and demonstrates that sequence features regulate this recruitment. We estimate the processivity of scanning ribosomes as they traverse the five prime UTR and show that the repressive effects of upstream open reading frames depend on the efficiency of both translation initiation and termination. Finally we determine the optimal initiation contexts by directly estimating the conversion of scanning to elongating ribosomes and demonstrate specific regulation of translation initiation at the endoplasmic reticulum. Our results open for the possibility of deconvoluting translation initiation into separate stages and provides the first view of global occupancy of ribosomal small subunits in a vertebrate. ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 07 03     Shield 2 SAMEA5752546 Computational Biology Unit ENA FIRST PUBLIC:2020 03 27T17:04:59Z|ENA LAST UPDATE:2019 07 03T10:01:14Z|External Id:SAMEA5752546|INSDC center name:Computational Biology Unit|INSDC first public:2020 03 27T17:04:59Z|INSDC last update:2019 07 03T10:01:14Z|INSDC status:public|Submitter Id:9|common name:zebrafish|dev stage:Shield|sample name:9|scientific name:Danio rerio                 NextSeq 500 sequencing ena EXPERIMENT Computational Biology Unit 22 08 2019 11:57:43:424 14 Shield 2 LSU None RCP seq   OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   ERP116106 NextSeq 500 sequencing ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 08 22     9775297004.0 128622329.0 ena RUN Computational Biology Unit 22 08 2019 11:57:43:424 14 0:76 A:4750565405;C:2540992255;G:1698817649;T:784832634;N:89061 76       4750565405 2540992255 1698817649 784832634 89061 ERX3511277 ERS3556005 ERA2100634 Computational Biology Unit|European Nucleotide Archive Computational Biology Unit 1 0.74003   0.5616   0.99855   0.03607   76   B   usable mapping rate illumina nextseq unknown other unknown bulk unknown unknown   Unknown 2019-07-03 Gastrula Embryo Undetermined Embryo Imprecise
9749 9749 ERR3489861 ERX3511276 ERS3556005 ERP116106 PRJEB33323 Deconstructing the individual steps of vertebrate translation initiation ena-STUDY-Computational Biology Unit-03-07-2019-10:11:34:314-422 Other In eukaryotes the number of ribosomes synthesizing a given protein depends on how many are recruited to its mRNA their success in navigating its five prime untranslated region UTR and whether they recognize its start codon. Initiation of translation is a rate limiting step in protein synthesis and key to gene expression control 1 but despite this centrality it remains poorly understood 2 3. Here we introduce ribosome complex profiling RCP seq to capture the transcriptome wide occupancy of scanning initiating elongating and terminating ribosome complexes in a higher eukaryote. We track scanning and elongating ribosomes across all five prime UTRs in zebrafish which enable us to assess the individual regulatory contributions from the three stages of initiation: ribosome recruitment scanning of the five prime UTR and recognition of the start codon. Our data sheds light on small subunit recruitment to mRNAs presenting evidence for the threading model and demonstrates that sequence features regulate this recruitment. We estimate the processivity of scanning ribosomes as they traverse the five prime UTR and show that the repressive effects of upstream open reading frames depend on the efficiency of both translation initiation and termination. Finally we determine the optimal initiation contexts by directly estimating the conversion of scanning to elongating ribosomes and demonstrate specific regulation of translation initiation at the endoplasmic reticulum. Our results open for the possibility of deconvoluting translation initiation into separate stages and provides the first view of global occupancy of ribosomal small subunits in a vertebrate. ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 07 03     Shield 2 SAMEA5752546 Computational Biology Unit ENA FIRST PUBLIC:2020 03 27T17:04:59Z|ENA LAST UPDATE:2019 07 03T10:01:14Z|External Id:SAMEA5752546|INSDC center name:Computational Biology Unit|INSDC first public:2020 03 27T17:04:59Z|INSDC last update:2019 07 03T10:01:14Z|INSDC status:public|Submitter Id:9|common name:zebrafish|dev stage:Shield|sample name:9|scientific name:Danio rerio                 NextSeq 500 sequencing ena EXPERIMENT Computational Biology Unit 22 08 2019 11:57:43:424 13 Shield 2 SSU None RCP seq   OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   ERP116106 NextSeq 500 sequencing ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 08 22     8210103300.0 108027675.0 ena RUN Computational Biology Unit 22 08 2019 11:57:43:424 13 0:76 A:3300825043;C:2576709678;G:1707115198;T:625376255;N:77126 76       3300825043 2576709678 1707115198 625376255 77126 ERX3511276 ERS3556005 ERA2100634 Computational Biology Unit|European Nucleotide Archive Computational Biology Unit 1 0.2787   0.17583   0.99752   0.50171   76   B   usable mapping rate illumina nextseq unknown other unknown bulk unknown unknown   Unknown 2019-07-03 Gastrula Embryo Undetermined Embryo Imprecise
9750 9750 ERR3489860 ERX3511275 ERS3556004 ERP116106 PRJEB33323 Deconstructing the individual steps of vertebrate translation initiation ena-STUDY-Computational Biology Unit-03-07-2019-10:11:34:314-422 Other In eukaryotes the number of ribosomes synthesizing a given protein depends on how many are recruited to its mRNA their success in navigating its five prime untranslated region UTR and whether they recognize its start codon. Initiation of translation is a rate limiting step in protein synthesis and key to gene expression control 1 but despite this centrality it remains poorly understood 2 3. Here we introduce ribosome complex profiling RCP seq to capture the transcriptome wide occupancy of scanning initiating elongating and terminating ribosome complexes in a higher eukaryote. We track scanning and elongating ribosomes across all five prime UTRs in zebrafish which enable us to assess the individual regulatory contributions from the three stages of initiation: ribosome recruitment scanning of the five prime UTR and recognition of the start codon. Our data sheds light on small subunit recruitment to mRNAs presenting evidence for the threading model and demonstrates that sequence features regulate this recruitment. We estimate the processivity of scanning ribosomes as they traverse the five prime UTR and show that the repressive effects of upstream open reading frames depend on the efficiency of both translation initiation and termination. Finally we determine the optimal initiation contexts by directly estimating the conversion of scanning to elongating ribosomes and demonstrate specific regulation of translation initiation at the endoplasmic reticulum. Our results open for the possibility of deconvoluting translation initiation into separate stages and provides the first view of global occupancy of ribosomal small subunits in a vertebrate. ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 07 03     Shield 1 SAMEA5752545 Computational Biology Unit ENA FIRST PUBLIC:2020 03 27T17:04:59Z|ENA LAST UPDATE:2019 07 03T10:01:14Z|External Id:SAMEA5752545|INSDC center name:Computational Biology Unit|INSDC first public:2020 03 27T17:04:59Z|INSDC last update:2019 07 03T10:01:14Z|INSDC status:public|Submitter Id:8|common name:zebrafish|dev stage:Shield|sample name:8|scientific name:Danio rerio                 NextSeq 500 sequencing ena EXPERIMENT Computational Biology Unit 22 08 2019 11:57:43:424 12 Shield 1 LSU None RCP seq   OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   ERP116106 NextSeq 500 sequencing ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 08 22     2437679936.0 32074736.0 ena RUN Computational Biology Unit 22 08 2019 11:57:43:424 12 0:76 A:764355184;C:710492003;G:664031460;T:298777374;N:23915 76       764355184 710492003 664031460 298777374 23915 ERX3511275 ERS3556004 ERA2100634 Computational Biology Unit|European Nucleotide Archive Computational Biology Unit 1 0.0406   0.02417   0.99908   0.61299   76   B   usable mapping rate illumina nextseq unknown other unknown bulk unknown unknown   Unknown 2019-07-03 Gastrula Embryo Undetermined Embryo Imprecise
9751 9751 ERR3489859 ERX3511274 ERS3556004 ERP116106 PRJEB33323 Deconstructing the individual steps of vertebrate translation initiation ena-STUDY-Computational Biology Unit-03-07-2019-10:11:34:314-422 Other In eukaryotes the number of ribosomes synthesizing a given protein depends on how many are recruited to its mRNA their success in navigating its five prime untranslated region UTR and whether they recognize its start codon. Initiation of translation is a rate limiting step in protein synthesis and key to gene expression control 1 but despite this centrality it remains poorly understood 2 3. Here we introduce ribosome complex profiling RCP seq to capture the transcriptome wide occupancy of scanning initiating elongating and terminating ribosome complexes in a higher eukaryote. We track scanning and elongating ribosomes across all five prime UTRs in zebrafish which enable us to assess the individual regulatory contributions from the three stages of initiation: ribosome recruitment scanning of the five prime UTR and recognition of the start codon. Our data sheds light on small subunit recruitment to mRNAs presenting evidence for the threading model and demonstrates that sequence features regulate this recruitment. We estimate the processivity of scanning ribosomes as they traverse the five prime UTR and show that the repressive effects of upstream open reading frames depend on the efficiency of both translation initiation and termination. Finally we determine the optimal initiation contexts by directly estimating the conversion of scanning to elongating ribosomes and demonstrate specific regulation of translation initiation at the endoplasmic reticulum. Our results open for the possibility of deconvoluting translation initiation into separate stages and provides the first view of global occupancy of ribosomal small subunits in a vertebrate. ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 07 03     Shield 1 SAMEA5752545 Computational Biology Unit ENA FIRST PUBLIC:2020 03 27T17:04:59Z|ENA LAST UPDATE:2019 07 03T10:01:14Z|External Id:SAMEA5752545|INSDC center name:Computational Biology Unit|INSDC first public:2020 03 27T17:04:59Z|INSDC last update:2019 07 03T10:01:14Z|INSDC status:public|Submitter Id:8|common name:zebrafish|dev stage:Shield|sample name:8|scientific name:Danio rerio                 NextSeq 500 sequencing ena EXPERIMENT Computational Biology Unit 22 08 2019 11:57:43:424 11 Shield 1 SSU None RCP seq   OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   ERP116106 NextSeq 500 sequencing ENA FIRST PUBLIC:2020 03 27|ENA LAST UPDATE:2019 08 22     3157243376.0 41542676.0 ena RUN Computational Biology Unit 22 08 2019 11:57:43:424 11 0:76 A:1443205052;C:715251024;G:633421305;T:365333650;N:32345 76       1443205052 715251024 633421305 365333650 32345 ERX3511274 ERS3556004 ERA2100634 Computational Biology Unit|European Nucleotide Archive Computational Biology Unit 1 0.47643   0.27944   0.99896   0.11464   76   B   usable mapping rate illumina nextseq unknown other unknown bulk unknown unknown   Unknown 2019-07-03 Gastrula Embryo Undetermined Embryo Imprecise
25295 25295 SRR25764045 SRX21486723 SRS18719024 SRP457105 PRJNA1009809 Dynamics of the zebrafish tRNAome during the maternal to zygotic transition [Ribo Seq] GSE241753 Other Time course analysis of tRNA abundance during zebrafish early embryonic development. Overall design: Wild type TLAB strain zebrafish embryos were grown in standard housing conditions.Unfertilized eggs 0 hpf were collected or embryos were staged and collected at consecutive developmental time points namelyat the 256 cell 2.5 hpf 1000 cell 3 hpf sphere 4 hpf shield 6 hpf and bud 10 hpf stages. Eggs and embryos were either flash frozen in liquid nitrogen or immediately processed. Samples were used for western blotting analysis polysome profiling ribosome profiling or mRNA and tRNA sequencing for investigating the regulation of tRNA gene expression and translational status during early zebrafish embryogenesis. parent bioproject:PRJNA1009800 pubmed:39402326   WT bud 10 hpf Ribo seq rep1 GSM7734770   source name:Gastrula|strain:TLAB strain|tissue:Gastrula|developmental stage:Bud 10 hpf|treatment:100 µg/ml CHX 100 µg/ml TIG|geo loc name:missing|collection date:missing WT bud 10 hpf Ribo seq rep1 We identified the A site location in each mapped read with Scikit ribo [Fang et al. 2018] which uses a random forest with recursive feature selection and a generalized linear model for accurate A site prediction based on matched ribosome profiling and RNA Seq datasets. Kallisto 0.44.0 with parameters b 100 single l 180 s 20 t 40 was used to quantify transcript abundances in Transcripts Per Million TPM from RNA Seq data based on the reference set of MANE annotated transcripts see Codon usage analysis for description of this annotation. To avoid memory errors due to the large size of the human genome and the presence of multiple transcript isoforms all RNAfold dependencies in Scikit ribo were omitted and the index was built separately for each chromosome. To make the hg38 GTF compatible with Scikit ribo transcript/UTR annotations were removed. For each transcript the start codon in the first exon and the stop codon in the last exon were adjusted to represent transcript start and end coordinates taking into account the gene strand. To estimate codon dwell times short 20 23 nt and long 28 33 nt ribosome footprints were analyzed separately. rRNA filtered reads were aligned to GRCz11.108 using STAR v2.6.1c [Dobin et al. 2013] the following options: outFilterMultimapNmax 1 seedSearchStartLmax 15 outSAMtype BAM SortedByCoordinate outFilterMismatchNmax 2 alignEndsType EndToEnd quantMode TranscriptomeSAM outSAMattributes NH HI AS nM NM MD We identified the A site location in each mapped read with Scikit ribo [Fang et al. 2018] which uses a random forest with recursive feature selection and a generalized linear model for accurate A site prediction based on matched ribosome profiling and RNA Seq datasets. Kallisto 0.44.0 with parameters b 100 single l 180 s 20 t 40 was used to quantify transcript abundances in Transcripts Per Million TPM from RNA Seq data based on the reference set of MANE annotated transcripts see Codon usage analysis for description of this annotation. To avoid memory err… Gastrula unperturbed growth conditions in E3 medium for zebrafish embryos. 200 whole embryos were flash frozen in liquid nitrogen and subsequently lysed in footprint lysis buffer containing 100 µg/ml CHX and 100 µg/ml TIG 0.1% NP 40 10 µg/ml aprotinin 20 µM leupeptin 2.5 µM pepstatin A 0.5 mM AEBSF and 1x Phosphatase Inhibitor Cocktail. Samples were vortexed vigorously triturated through a 26G gauge needle and spun down for 7 minutes at 16 000xg/ 4°C. Supernatant was transferred to a new tube. 20 µg RNA in 200 µl polysome lysis buffer were digested with 50 U RNase I for 45 minutes at 2 000 rpm/22°C. post incubation on ice for 5 minutes extracts were pre cleared by centrifugation for 5 minutes at 3 000 g/ 4°C. Ribosomes were pelleted through 3 ml of a sucrose cushion 1 M sucrose 20 mM Tris pH=8.0 140 mM KCl 5 mM MgCl2 1 mM DTT by spinning the layered solutions in the Type 70 Ti rotor for 120 minutes at 50 000 rpm/ 4°C. Ribosome pellets were rinsed once dissolved in 200 µl drug free polysome lysis buffer and incubated with 200 U hiPSC or 300 U NPC RNase I for 45 minutes at 2 000 rpm/22°C. Ribosome footprint libraries were prepared essentially as described McGlincy and Ingolia 2017; Wu 2019 with minor modifications. RNase I digestion was stopped by addition of 100 U Superase In and extracts were loaded on a sucrose cushion. The pellet was dissolved in 400 µl LiDS/LET lysis buffer and RNA was extracted with the acid phenol protocol. Fragments in the range of 19 to 32 nucleotides were isolated by gel size selection with T4 PNK and ligated to pre adenylated adapters containing 5 random nucleotides at their 5’ ends McGlinzy 2017 with T4 RNA Ligase 2 truncated KQ. Adapter ligated RNA was subjected to rRNA depletion using the Ribo Seq riboPOOL h/m/r depletion kit siTOOLs for CHX only samples and legacy RiboZero Gold kit Illumina for CHX+TIG samples The rRNA depleted footprints were reverse transcribed with Protoscript II and cDNA was circularized with recombinant TS2126 RNA ligase 1 commercially available as CircLigase. Libraries were constructed from circularized cDNA with KAP… Embryos were grown in standard housing conditions namely28°C at a 14/10 hour light/dark cycle. strain:TLAB strain|tissue:Gastrula|developmental stage:Bud 10 hpf|treatment:100 µg/ml CHX 100 µg/ml TIG GSM7734770 GSM7734770: WT bud 10 hpf Ribo seq rep1; Danio rerio; OTHER GSM7734770 r1 GSM7734770 1 200 whole embryos were flash frozen in liquid nitrogen and subsequently lysed in footprint lysis buffer containing 100 µg/ml CHX and 100 µg/ml TIG 0.1% NP 40 10 µg/ml aprotinin 20 µM leupeptin 2.5 µM pepstatin A 0.5 mM AEBSF and 1x Phosphatase Inhibitor Cocktail. Samples were vortexed vigorously triturated through a 26G gauge needle and spun down for 7 minutes at 16 000xg/ 4°C. Supernatant was transferred to a new tube. 20 µg RNA in 200 µl polysome lysis buffer were digested with 50 U RNase I for 45 minutes at 2 000 rpm/22°C. post incubation on ice for 5 minutes extracts were pre cleared by centrifugation for 5 minutes at 3 000 g/ 4°C. Ribosomes were pelleted through 3 ml of a sucrose cushion 1 M sucrose 20 mM Tris pH=8.0 140 mM KCl 5 mM MgCl2 1 mM DTT by spinning the layered solutions in the Type 70 Ti rotor for 120 minutes at 50 000 rpm/ 4°C. Ribosome pellets were rinsed once dissolved in 200 µl drug free polysome lysis buffer and incubated with 200 U hiPSC or 300 U NPC RNase I for 45 minutes at 2 000 rpm/22°C. Ribosome footprint libraries were prepared essentially as described McGlincy and Ingolia 2017; Wu 2019 with minor modifications. RNase I digestion was stopped by addition of 100 U Superase In and extracts were loaded on a sucrose cushion. The pellet was dissolved in 400 µl LiDS/LET lysis buffer and RNA was extracted with the acid phenol protocol. Fragments in the range of 19 to 32 nucleotides were isolated by gel size selection with T4 PNK and ligated to pre adenylated adapters containing 5 random nucleotides at their five prime ends McGlinzy 2017 with T4 RNA Ligase 2 truncated KQ. Adapter ligated RNA was subjected to rRNA depletion using the Ribo Seq riboPOOL h/m/r depletion kit siTOOLs for CHX only samples and legacy RiboZero Gold kit Illumina for CHX+TIG samples The rRNA depleted footprints were reverse transcribed with Protoscript II and cDNA was circularized with recombinant TS2126 RNA ligase 1 commercially available as CircLigase. Libraries were constructed from circularized cDNA …   OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA NextSeq 500   SRP457105     WT_ribo_bud_1.fastq.gz fastq 1476985703.0 55745002.0 GSM7734770 r1 0:26.50 A:266369385;C:466461341;G:473469631;T:270671671;N:13675 26       266369385 466461341 473469631 270671671 13675 SRX21486723 SRS18719024 SRA1700409 Mechanisms of Protein Biogenesis, Max Planck Institute for Biochemistry Max Planck Institute of Biochemistry 1 0.773   0.14121   0.82242   0.78464   30   B   usable mapping rate illumina nextseq 5prime rrna_depletion ribozero bulk unknown unknown   Germany 2023-08-28 Gastrula Embryo Whole Organism All anatomical structures
31943 31943 SRR28790250 SRX24354554 SRS21112322 SRP503769 PRJNA1104180 Double stranded RNA triggers a distinct integrated stress response in the early embryo [RNA seq Ribo seq] GSE265771 Other Double stranded RNA dsRNA is associated with virus infections and is present as by products during the transcription of synthetic mRNA which has been widely used in gene gain of function studies and serves as a core component in emerging mRNA based therapies1 4. The presence of dsRNA in host cells induces an integrated stress response that functions to prevent virus replication and infection5 6. Unlike differentiated cells undifferentiated cells adopt a distinct defense strategy against RNA virus infection7 but the mechanism is unclear. We show a previously unidentified response triggered by dsRNA in the early embryo. Although dsRNA causes a global protein translation inhibition in a PKR eIF2a independent manner and leads to developmental delay and cell necrosis it also strongly induces p53 activation which then upregulates Interferon Stimulated Genes independently of interferon ligands. Importantly we demonstrate that the burst of p53 signaling dose not result in cell death but functions as a protective mechanism against deleterious translation blockage by slowing down global protein degradation via ISGylation. Our work has identified a distinct dsRNA induced stress response in the embryo reflecting an ancient innate immune memory before the establishment of the IFN system. It also raises the provocative question as to the original protective role of p53 during evolution. Overall design: To characterize the distribution of ribosomes on mRNA we performed Ribosome profiling Ribo seq analysis with RNA seq on dsRNA injected embryos.       Zebrafish Riboseq high dose dsRNA2 GSM8228804   source name:whole embryo|tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:dsRNA|geo loc name:missing|collection date:missing Zebrafish Riboseq high dose dsRNA2 The library underwent quality control assessment and was subjected to Illumina Novaseq 6000 sequencing. Bio informatics analysis of RIBOseq profilling was performed to analyze the library data. Assembly: GRCz10 Supplementary files format and content: Excel file includes raw counts for each Sample Library strategy: Ribo seq whole embryo   Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components including undigested material such as cell nuclei.Then RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA while the RNA fragments protected by ribosomes were preserved. Subsequently an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx and the corresponding bands were collected. The recovered bands were subjected to ligation reaction where a preadenylylated and 3’ blocked linker 5’ rApp CTGTAGGCACCATCAAT NH2 3’ was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre CL4111K resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR and barcode sequences were incorporated.   tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:dsRNA GSM8228804 GSM8228804: Zebrafish Riboseq high dose dsRNA2; Danio rerio; OTHER GSM8228804 r1 GSM8228804 1 Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components including undigested material such as cell nuclei.Then RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA while the RNA fragments protected by ribosomes were preserved. Subsequently an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx and the corresponding bands were collected. The recovered bands were subjected to ligation reaction where a preadenylylated and three prime blocked linker five prime rApp CTGTAGGCACCATCAAT NH2 three prime was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre CL4111K resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR and barcode sequences were incorporated.   OTHER TRANSCRIPTOMIC other PAIRED ILLUMINA Illumina NovaSeq 6000   SRP503769     dsR_2.R1.raw.fastq.gz dsR_2.R2.raw.fastq.gz fastq fastq 15877367026.0 52574063.0 GSM8228804 r1 0:151 1:151 A:3017944891;C:2902425937;G:7224440180;T:2731881397;N:674621 151 151     3017944891 2902425937 7224440180 2731881397 674621 SRX24354554 SRS21112322 SRA1852128 ShanDong University ShanDong University                       T T mates < 9% mapping rate illumina novaseq_era 3prime random_priming unknown bulk unknown unknown   China 2024-04-24 Gastrula Embryo Whole Organism All anatomical structures
31944 31944 SRR28790251 SRX24354553 SRS21112321 SRP503769 PRJNA1104180 Double stranded RNA triggers a distinct integrated stress response in the early embryo [RNA seq Ribo seq] GSE265771 Other Double stranded RNA dsRNA is associated with virus infections and is present as by products during the transcription of synthetic mRNA which has been widely used in gene gain of function studies and serves as a core component in emerging mRNA based therapies1 4. The presence of dsRNA in host cells induces an integrated stress response that functions to prevent virus replication and infection5 6. Unlike differentiated cells undifferentiated cells adopt a distinct defense strategy against RNA virus infection7 but the mechanism is unclear. We show a previously unidentified response triggered by dsRNA in the early embryo. Although dsRNA causes a global protein translation inhibition in a PKR eIF2a independent manner and leads to developmental delay and cell necrosis it also strongly induces p53 activation which then upregulates Interferon Stimulated Genes independently of interferon ligands. Importantly we demonstrate that the burst of p53 signaling dose not result in cell death but functions as a protective mechanism against deleterious translation blockage by slowing down global protein degradation via ISGylation. Our work has identified a distinct dsRNA induced stress response in the embryo reflecting an ancient innate immune memory before the establishment of the IFN system. It also raises the provocative question as to the original protective role of p53 during evolution. Overall design: To characterize the distribution of ribosomes on mRNA we performed Ribosome profiling Ribo seq analysis with RNA seq on dsRNA injected embryos.       Zebrafish Riboseq high dose dsRNA1 GSM8228803   source name:whole embryo|tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:dsRNA|geo loc name:missing|collection date:missing Zebrafish Riboseq high dose dsRNA1 The library underwent quality control assessment and was subjected to Illumina Novaseq 6000 sequencing. Bio informatics analysis of RIBOseq profilling was performed to analyze the library data. Assembly: GRCz10 Supplementary files format and content: Excel file includes raw counts for each Sample Library strategy: Ribo seq whole embryo   Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components including undigested material such as cell nuclei.Then RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA while the RNA fragments protected by ribosomes were preserved. Subsequently an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx and the corresponding bands were collected. The recovered bands were subjected to ligation reaction where a preadenylylated and 3’ blocked linker 5’ rApp CTGTAGGCACCATCAAT NH2 3’ was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre CL4111K resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR and barcode sequences were incorporated.   tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:dsRNA GSM8228803 GSM8228803: Zebrafish Riboseq high dose dsRNA1; Danio rerio; OTHER GSM8228803 r1 GSM8228803 1 Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components including undigested material such as cell nuclei.Then RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA while the RNA fragments protected by ribosomes were preserved. Subsequently an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx and the corresponding bands were collected. The recovered bands were subjected to ligation reaction where a preadenylylated and three prime blocked linker five prime rApp CTGTAGGCACCATCAAT NH2 three prime was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre CL4111K resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR and barcode sequences were incorporated.   OTHER TRANSCRIPTOMIC other PAIRED ILLUMINA Illumina NovaSeq 6000   SRP503769     dsR_1.R1.raw.fastq.gz dsR_1.R2.raw.fastq.gz fastq fastq 13027928774.0 43138837.0 GSM8228803 r1 0:151 1:151 A:2538099201;C:2400262316;G:5852047273;T:2236961902;N:558082 151 151     2538099201 2400262316 5852047273 2236961902 558082 SRX24354553 SRS21112321 SRA1852128 ShanDong University ShanDong University                       T T mates < 9% mapping rate illumina novaseq_era 3prime random_priming unknown bulk unknown unknown   China 2024-04-24 Gastrula Embryo Whole Organism All anatomical structures
31945 31945 SRR28790252 SRX24354552 SRS21112320 SRP503769 PRJNA1104180 Double stranded RNA triggers a distinct integrated stress response in the early embryo [RNA seq Ribo seq] GSE265771 Other Double stranded RNA dsRNA is associated with virus infections and is present as by products during the transcription of synthetic mRNA which has been widely used in gene gain of function studies and serves as a core component in emerging mRNA based therapies1 4. The presence of dsRNA in host cells induces an integrated stress response that functions to prevent virus replication and infection5 6. Unlike differentiated cells undifferentiated cells adopt a distinct defense strategy against RNA virus infection7 but the mechanism is unclear. We show a previously unidentified response triggered by dsRNA in the early embryo. Although dsRNA causes a global protein translation inhibition in a PKR eIF2a independent manner and leads to developmental delay and cell necrosis it also strongly induces p53 activation which then upregulates Interferon Stimulated Genes independently of interferon ligands. Importantly we demonstrate that the burst of p53 signaling dose not result in cell death but functions as a protective mechanism against deleterious translation blockage by slowing down global protein degradation via ISGylation. Our work has identified a distinct dsRNA induced stress response in the embryo reflecting an ancient innate immune memory before the establishment of the IFN system. It also raises the provocative question as to the original protective role of p53 during evolution. Overall design: To characterize the distribution of ribosomes on mRNA we performed Ribosome profiling Ribo seq analysis with RNA seq on dsRNA injected embryos.       Zebrafish Riboseq high dose uninj2 GSM8228802   source name:whole embryo|tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:uninjected|geo loc name:missing|collection date:missing Zebrafish Riboseq high dose uninj2 The library underwent quality control assessment and was subjected to Illumina Novaseq 6000 sequencing. Bio informatics analysis of RIBOseq profilling was performed to analyze the library data. Assembly: GRCz10 Supplementary files format and content: Excel file includes raw counts for each Sample Library strategy: Ribo seq whole embryo   Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components including undigested material such as cell nuclei.Then RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA while the RNA fragments protected by ribosomes were preserved. Subsequently an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx and the corresponding bands were collected. The recovered bands were subjected to ligation reaction where a preadenylylated and 3’ blocked linker 5’ rApp CTGTAGGCACCATCAAT NH2 3’ was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre CL4111K resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR and barcode sequences were incorporated.   tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:uninjected GSM8228802 GSM8228802: Zebrafish Riboseq high dose uninj2; Danio rerio; OTHER GSM8228802 r1 GSM8228802 1 Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components including undigested material such as cell nuclei.Then RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA while the RNA fragments protected by ribosomes were preserved. Subsequently an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx and the corresponding bands were collected. The recovered bands were subjected to ligation reaction where a preadenylylated and three prime blocked linker five prime rApp CTGTAGGCACCATCAAT NH2 three prime was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre CL4111K resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR and barcode sequences were incorporated.   OTHER TRANSCRIPTOMIC other PAIRED ILLUMINA Illumina NovaSeq 6000   SRP503769     un_2.R1.raw.fastq.gz un_2.R2.raw.fastq.gz fastq fastq 13637060056.0 45155828.0 GSM8228802 r1 0:151 1:151 A:2596015831;C:2388366938;G:6201010197;T:2451085273;N:581817 151 151     2596015831 2388366938 6201010197 2451085273 581817 SRX24354552 SRS21112320 SRA1852128 ShanDong University ShanDong University                       T T mates < 9% mapping rate illumina novaseq_era 3prime random_priming unknown bulk unknown unknown   China 2024-04-24 Gastrula Embryo Whole Organism All anatomical structures
31946 31946 SRR28790253 SRX24354551 SRS21112319 SRP503769 PRJNA1104180 Double stranded RNA triggers a distinct integrated stress response in the early embryo [RNA seq Ribo seq] GSE265771 Other Double stranded RNA dsRNA is associated with virus infections and is present as by products during the transcription of synthetic mRNA which has been widely used in gene gain of function studies and serves as a core component in emerging mRNA based therapies1 4. The presence of dsRNA in host cells induces an integrated stress response that functions to prevent virus replication and infection5 6. Unlike differentiated cells undifferentiated cells adopt a distinct defense strategy against RNA virus infection7 but the mechanism is unclear. We show a previously unidentified response triggered by dsRNA in the early embryo. Although dsRNA causes a global protein translation inhibition in a PKR eIF2a independent manner and leads to developmental delay and cell necrosis it also strongly induces p53 activation which then upregulates Interferon Stimulated Genes independently of interferon ligands. Importantly we demonstrate that the burst of p53 signaling dose not result in cell death but functions as a protective mechanism against deleterious translation blockage by slowing down global protein degradation via ISGylation. Our work has identified a distinct dsRNA induced stress response in the embryo reflecting an ancient innate immune memory before the establishment of the IFN system. It also raises the provocative question as to the original protective role of p53 during evolution. Overall design: To characterize the distribution of ribosomes on mRNA we performed Ribosome profiling Ribo seq analysis with RNA seq on dsRNA injected embryos.       Zebrafish Riboseq high dose uninj1 GSM8228801   source name:whole embryo|tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:uninjected|geo loc name:missing|collection date:missing Zebrafish Riboseq high dose uninj1 The library underwent quality control assessment and was subjected to Illumina Novaseq 6000 sequencing. Bio informatics analysis of RIBOseq profilling was performed to analyze the library data. Assembly: GRCz10 Supplementary files format and content: Excel file includes raw counts for each Sample Library strategy: Ribo seq whole embryo   Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components including undigested material such as cell nuclei.Then RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA while the RNA fragments protected by ribosomes were preserved. Subsequently an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx and the corresponding bands were collected. The recovered bands were subjected to ligation reaction where a preadenylylated and 3’ blocked linker 5’ rApp CTGTAGGCACCATCAAT NH2 3’ was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre CL4111K resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR and barcode sequences were incorporated.   tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:uninjected GSM8228801 GSM8228801: Zebrafish Riboseq high dose uninj1; Danio rerio; OTHER GSM8228801 r1 GSM8228801 1 Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components including undigested material such as cell nuclei.Then RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA while the RNA fragments protected by ribosomes were preserved. Subsequently an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx and the corresponding bands were collected. The recovered bands were subjected to ligation reaction where a preadenylylated and three prime blocked linker five prime rApp CTGTAGGCACCATCAAT NH2 three prime was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre CL4111K resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR and barcode sequences were incorporated.   OTHER TRANSCRIPTOMIC other PAIRED ILLUMINA Illumina NovaSeq 6000   SRP503769     un_1.R1.raw.fastq.gz un_1.R2.raw.fastq.gz fastq fastq 13499333560.0 44699780.0 GSM8228801 r1 0:151 1:151 A:2587725963;C:2439789499;G:6080777505;T:2390460459;N:580134 151 151     2587725963 2439789499 6080777505 2390460459 580134 SRX24354551 SRS21112319 SRA1852128 ShanDong University ShanDong University                       T T mates < 9% mapping rate illumina novaseq_era 3prime random_priming unknown bulk unknown unknown   China 2024-04-24 Gastrula Embryo Whole Organism All anatomical structures
31947 31947 SRR28790254 SRX24354550 SRS21112318 SRP503769 PRJNA1104180 Double stranded RNA triggers a distinct integrated stress response in the early embryo [RNA seq Ribo seq] GSE265771 Other Double stranded RNA dsRNA is associated with virus infections and is present as by products during the transcription of synthetic mRNA which has been widely used in gene gain of function studies and serves as a core component in emerging mRNA based therapies1 4. The presence of dsRNA in host cells induces an integrated stress response that functions to prevent virus replication and infection5 6. Unlike differentiated cells undifferentiated cells adopt a distinct defense strategy against RNA virus infection7 but the mechanism is unclear. We show a previously unidentified response triggered by dsRNA in the early embryo. Although dsRNA causes a global protein translation inhibition in a PKR eIF2a independent manner and leads to developmental delay and cell necrosis it also strongly induces p53 activation which then upregulates Interferon Stimulated Genes independently of interferon ligands. Importantly we demonstrate that the burst of p53 signaling dose not result in cell death but functions as a protective mechanism against deleterious translation blockage by slowing down global protein degradation via ISGylation. Our work has identified a distinct dsRNA induced stress response in the embryo reflecting an ancient innate immune memory before the establishment of the IFN system. It also raises the provocative question as to the original protective role of p53 during evolution. Overall design: To characterize the distribution of ribosomes on mRNA we performed Ribosome profiling Ribo seq analysis with RNA seq on dsRNA injected embryos.       Zebrafish Riboseq low dose dsRNA2 GSM8228800   source name:whole embryo|tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:dsRNA|geo loc name:missing|collection date:missing Zebrafish Riboseq low dose dsRNA2 The library underwent quality control assessment and was subjected to Illumina Novaseq 6000 sequencing. Bio informatics analysis of RIBOseq profilling was performed to analyze the library data. Assembly: GRCz10 Supplementary files format and content: Excel file includes raw counts for each Sample Library strategy: Ribo seq whole embryo   Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components including undigested material such as cell nuclei.Then RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA while the RNA fragments protected by ribosomes were preserved. Subsequently an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx and the corresponding bands were collected. The recovered bands were subjected to ligation reaction where a preadenylylated and 3’ blocked linker 5’ rApp CTGTAGGCACCATCAAT NH2 3’ was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre CL4111K resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR and barcode sequences were incorporated.   tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:dsRNA GSM8228800 GSM8228800: Zebrafish Riboseq low dose dsRNA2; Danio rerio; OTHER GSM8228800 r1 GSM8228800 1 Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components including undigested material such as cell nuclei.Then RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA while the RNA fragments protected by ribosomes were preserved. Subsequently an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx and the corresponding bands were collected. The recovered bands were subjected to ligation reaction where a preadenylylated and three prime blocked linker five prime rApp CTGTAGGCACCATCAAT NH2 three prime was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre CL4111K resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR and barcode sequences were incorporated.   OTHER TRANSCRIPTOMIC other PAIRED ILLUMINA Illumina NovaSeq 6000   SRP503769     dsR_2.raw.1.fastq.gz dsR_2.raw.2.fastq.gz fastq fastq 13335544766.0 44157433.0 GSM8228800 r1 0:151 1:151 A:2383528724;C:2262795235;G:6593680813;T:2094730217;N:809777 151 151     2383528724 2262795235 6593680813 2094730217 809777 SRX24354550 SRS21112318 SRA1852128 ShanDong University ShanDong University                       T T mates < 9% mapping rate illumina novaseq_era 3prime random_priming unknown bulk unknown unknown   China 2024-04-24 Gastrula Embryo Whole Organism All anatomical structures
31948 31948 SRR28790255 SRX24354549 SRS21112317 SRP503769 PRJNA1104180 Double stranded RNA triggers a distinct integrated stress response in the early embryo [RNA seq Ribo seq] GSE265771 Other Double stranded RNA dsRNA is associated with virus infections and is present as by products during the transcription of synthetic mRNA which has been widely used in gene gain of function studies and serves as a core component in emerging mRNA based therapies1 4. The presence of dsRNA in host cells induces an integrated stress response that functions to prevent virus replication and infection5 6. Unlike differentiated cells undifferentiated cells adopt a distinct defense strategy against RNA virus infection7 but the mechanism is unclear. We show a previously unidentified response triggered by dsRNA in the early embryo. Although dsRNA causes a global protein translation inhibition in a PKR eIF2a independent manner and leads to developmental delay and cell necrosis it also strongly induces p53 activation which then upregulates Interferon Stimulated Genes independently of interferon ligands. Importantly we demonstrate that the burst of p53 signaling dose not result in cell death but functions as a protective mechanism against deleterious translation blockage by slowing down global protein degradation via ISGylation. Our work has identified a distinct dsRNA induced stress response in the embryo reflecting an ancient innate immune memory before the establishment of the IFN system. It also raises the provocative question as to the original protective role of p53 during evolution. Overall design: To characterize the distribution of ribosomes on mRNA we performed Ribosome profiling Ribo seq analysis with RNA seq on dsRNA injected embryos.       Zebrafish Riboseq low dose dsRNA1 GSM8228799   source name:whole embryo|tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:dsRNA|geo loc name:missing|collection date:missing Zebrafish Riboseq low dose dsRNA1 The library underwent quality control assessment and was subjected to Illumina Novaseq 6000 sequencing. Bio informatics analysis of RIBOseq profilling was performed to analyze the library data. Assembly: GRCz10 Supplementary files format and content: Excel file includes raw counts for each Sample Library strategy: Ribo seq whole embryo   Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components including undigested material such as cell nuclei.Then RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA while the RNA fragments protected by ribosomes were preserved. Subsequently an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx and the corresponding bands were collected. The recovered bands were subjected to ligation reaction where a preadenylylated and 3’ blocked linker 5’ rApp CTGTAGGCACCATCAAT NH2 3’ was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre CL4111K resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR and barcode sequences were incorporated.   tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:dsRNA GSM8228799 GSM8228799: Zebrafish Riboseq low dose dsRNA1; Danio rerio; OTHER GSM8228799 r1 GSM8228799 1 Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components including undigested material such as cell nuclei.Then RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA while the RNA fragments protected by ribosomes were preserved. Subsequently an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx and the corresponding bands were collected. The recovered bands were subjected to ligation reaction where a preadenylylated and three prime blocked linker five prime rApp CTGTAGGCACCATCAAT NH2 three prime was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre CL4111K resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR and barcode sequences were incorporated.   OTHER TRANSCRIPTOMIC other PAIRED ILLUMINA Illumina NovaSeq 6000   SRP503769     dsR_1.raw.1.fastq.gz dsR_1.raw.2.fastq.gz fastq fastq 12530433302.0 41491501.0 GSM8228799 r1 0:151 1:151 A:2300207705;C:2164355626;G:6050556625;T:2014558360;N:754986 151 151     2300207705 2164355626 6050556625 2014558360 754986 SRX24354549 SRS21112317 SRA1852128 ShanDong University ShanDong University                       T T mates < 9% mapping rate illumina novaseq_era 3prime random_priming unknown bulk unknown unknown   China 2024-04-24 Gastrula Embryo Whole Organism All anatomical structures
31949 31949 SRR28790256 SRX24354548 SRS21112316 SRP503769 PRJNA1104180 Double stranded RNA triggers a distinct integrated stress response in the early embryo [RNA seq Ribo seq] GSE265771 Other Double stranded RNA dsRNA is associated with virus infections and is present as by products during the transcription of synthetic mRNA which has been widely used in gene gain of function studies and serves as a core component in emerging mRNA based therapies1 4. The presence of dsRNA in host cells induces an integrated stress response that functions to prevent virus replication and infection5 6. Unlike differentiated cells undifferentiated cells adopt a distinct defense strategy against RNA virus infection7 but the mechanism is unclear. We show a previously unidentified response triggered by dsRNA in the early embryo. Although dsRNA causes a global protein translation inhibition in a PKR eIF2a independent manner and leads to developmental delay and cell necrosis it also strongly induces p53 activation which then upregulates Interferon Stimulated Genes independently of interferon ligands. Importantly we demonstrate that the burst of p53 signaling dose not result in cell death but functions as a protective mechanism against deleterious translation blockage by slowing down global protein degradation via ISGylation. Our work has identified a distinct dsRNA induced stress response in the embryo reflecting an ancient innate immune memory before the establishment of the IFN system. It also raises the provocative question as to the original protective role of p53 during evolution. Overall design: To characterize the distribution of ribosomes on mRNA we performed Ribosome profiling Ribo seq analysis with RNA seq on dsRNA injected embryos.       Zebrafish Riboseq low dose uninj2 GSM8228798   source name:whole embryo|tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:uninjected|geo loc name:missing|collection date:missing Zebrafish Riboseq low dose uninj2 The library underwent quality control assessment and was subjected to Illumina Novaseq 6000 sequencing. Bio informatics analysis of RIBOseq profilling was performed to analyze the library data. Assembly: GRCz10 Supplementary files format and content: Excel file includes raw counts for each Sample Library strategy: Ribo seq whole embryo   Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components including undigested material such as cell nuclei.Then RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA while the RNA fragments protected by ribosomes were preserved. Subsequently an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx and the corresponding bands were collected. The recovered bands were subjected to ligation reaction where a preadenylylated and 3’ blocked linker 5’ rApp CTGTAGGCACCATCAAT NH2 3’ was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre CL4111K resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR and barcode sequences were incorporated.   tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:uninjected GSM8228798 GSM8228798: Zebrafish Riboseq low dose uninj2; Danio rerio; OTHER GSM8228798 r1 GSM8228798 1 Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components including undigested material such as cell nuclei.Then RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA while the RNA fragments protected by ribosomes were preserved. Subsequently an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx and the corresponding bands were collected. The recovered bands were subjected to ligation reaction where a preadenylylated and three prime blocked linker five prime rApp CTGTAGGCACCATCAAT NH2 three prime was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre CL4111K resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR and barcode sequences were incorporated.   OTHER TRANSCRIPTOMIC other PAIRED ILLUMINA Illumina NovaSeq 6000   SRP503769     un_2.raw.1.fastq.gz un_2.raw.2.fastq.gz fastq fastq 12132018896.0 40172248.0 GSM8228798 r1 0:151 1:151 A:2225485331;C:2027579165;G:5953072417;T:1925141784;N:740199 151 151     2225485331 2027579165 5953072417 1925141784 740199 SRX24354548 SRS21112316 SRA1852128 ShanDong University ShanDong University                       T T mates < 9% mapping rate illumina novaseq_era 3prime random_priming unknown bulk unknown unknown   China 2024-04-24 Gastrula Embryo Whole Organism All anatomical structures
31950 31950 SRR28790257 SRX24354547 SRS21112315 SRP503769 PRJNA1104180 Double stranded RNA triggers a distinct integrated stress response in the early embryo [RNA seq Ribo seq] GSE265771 Other Double stranded RNA dsRNA is associated with virus infections and is present as by products during the transcription of synthetic mRNA which has been widely used in gene gain of function studies and serves as a core component in emerging mRNA based therapies1 4. The presence of dsRNA in host cells induces an integrated stress response that functions to prevent virus replication and infection5 6. Unlike differentiated cells undifferentiated cells adopt a distinct defense strategy against RNA virus infection7 but the mechanism is unclear. We show a previously unidentified response triggered by dsRNA in the early embryo. Although dsRNA causes a global protein translation inhibition in a PKR eIF2a independent manner and leads to developmental delay and cell necrosis it also strongly induces p53 activation which then upregulates Interferon Stimulated Genes independently of interferon ligands. Importantly we demonstrate that the burst of p53 signaling dose not result in cell death but functions as a protective mechanism against deleterious translation blockage by slowing down global protein degradation via ISGylation. Our work has identified a distinct dsRNA induced stress response in the embryo reflecting an ancient innate immune memory before the establishment of the IFN system. It also raises the provocative question as to the original protective role of p53 during evolution. Overall design: To characterize the distribution of ribosomes on mRNA we performed Ribosome profiling Ribo seq analysis with RNA seq on dsRNA injected embryos.       Zebrafish Riboseq low dose uninj1 GSM8228797   source name:whole embryo|tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:uninjected|geo loc name:missing|collection date:missing Zebrafish Riboseq low dose uninj1 The library underwent quality control assessment and was subjected to Illumina Novaseq 6000 sequencing. Bio informatics analysis of RIBOseq profilling was performed to analyze the library data. Assembly: GRCz10 Supplementary files format and content: Excel file includes raw counts for each Sample Library strategy: Ribo seq whole embryo   Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components including undigested material such as cell nuclei.Then RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA while the RNA fragments protected by ribosomes were preserved. Subsequently an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx and the corresponding bands were collected. The recovered bands were subjected to ligation reaction where a preadenylylated and 3’ blocked linker 5’ rApp CTGTAGGCACCATCAAT NH2 3’ was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre CL4111K resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR and barcode sequences were incorporated.   tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:uninjected GSM8228797 GSM8228797: Zebrafish Riboseq low dose uninj1; Danio rerio; OTHER GSM8228797 r1 GSM8228797 1 Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components including undigested material such as cell nuclei.Then RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA while the RNA fragments protected by ribosomes were preserved. Subsequently an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx and the corresponding bands were collected. The recovered bands were subjected to ligation reaction where a preadenylylated and three prime blocked linker five prime rApp CTGTAGGCACCATCAAT NH2 three prime was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre CL4111K resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR and barcode sequences were incorporated.   OTHER TRANSCRIPTOMIC other PAIRED ILLUMINA Illumina NovaSeq 6000   SRP503769     un_1.raw.1.fastq.gz un_1.raw.2.fastq.gz fastq fastq 10423454802.0 34514751.0 GSM8228797 r1 0:151 1:151 A:1931094519;C:1788894999;G:5028533789;T:1674294221;N:637274 151 151     1931094519 1788894999 5028533789 1674294221 637274 SRX24354547 SRS21112315 SRA1852128 ShanDong University ShanDong University                       T T mates < 9% mapping rate illumina novaseq_era 3prime random_priming unknown bulk unknown unknown   China 2024-04-24 Gastrula Embryo Whole Organism All anatomical structures
41548 41548 SRR5017075 SRX2345570 SRS1796136 SRP093295 PRJNA353372 N6 methyladenosine dynamics during early vertebrate embryogenesis GSE89815 Other Early vertebrate embryogenesis is characterized by extensive post transcriptional regulation during the maternal to zygotic transition. The N6 methyladenosine m6A modifications on mRNA has been shown to affect both translation and stability of transcripts. Here we investigate the m6A topology during early vertebrate embryogenesis and its association with RNA stability translation efficiency and effect on miR 430 degradation kinetics. Notably we find a strong association of m6A with cytoplasmic polyadenylation and translational efficiency prior to zygotic genome activation. Genes required for zygotic genome activation such as nanog and pou5f3 display dynamic m6A levels. post zygotic genome activation m6A is associated with improved stability and dampens the effect of miR 430 mediated degradation. Through sequence analyses we identified enrichment of motifs for RNA binding proteins involved in translational regulation and RNA degradation. We propose a role for m6A in multiple mRNA regulatory mechanisms for the first time in an in vivo system and improve our understanding of the combinatorial code behind the complex post transcriptional regulation of reprogramming during early vertebrate development. Overall design: Examination of m6A in four different developmental stages       input shield rep2 GSM2390028   tissue:Embryos|developmental stage:Shield stage embryos|strain:AB wild type input shield rep2 Base calling The sequencing output from each of the 4 lanes the sequencing run was trimmed 7 nucleotides in the 5’end and de multiplexed 4 samples in each lane. Reads were mapped to Zv10 using the STAR aligner Dobin et al. 2013 with options seedSearchStartLmax 15 clip3pNbases 10 clip5pNbases 10 outFilterMultimapNmax 20 outFilterMismatchNoverLmax 0.05 outFilterMatchNminOverLread 0.0 outFilterMatchNmin 15 outFilterScoreMinOverLread 0.0. We performed peak calling and detection of differentially methylated genes using ExomePeak Meng et al. 2013. Genome build: Zv10 Supplementary files format and content: Bed files with enriched regions from each developmental stage and txt file with raw and normalized read counts for each gene using the input samples Embryos   We isolated total RNA from 4 stages 1 cell/20 minutes post fertilization 2 4 and 6 hpf in batches of 200 embryos using TRIzol Invitrogen cat.no. 15596 018. We added ERCC spike in RNA TermoFisher scientific cat.no. 4456740 to the trizol. For each sequencing library two batches of total RNA from 200 embryos were merged and enriched for polyA+ RNA using Dynabeads® mRNA Purification Kit Ambion #61006. The m6A RIP experiment was carried out as previously described Ke et al. 2015 and the protocol can be found in supplementary file 1. Briefly polyA+ enriched RNA was partially fragmented by alkaline hydrolysis ethanol precipitated and SDS PAGE size selected for 20 80nt. Part of the fragmented RNA was used as input and the rest was immunoprecipitated at 4oC for 2 h using Dynabeads Protein A Life Technologies # 10008D conjugated anti m6A antibody Synaptic systems # 202003. post stringent washing the bound RNA was eluted with 0.5mg/ml N6 methyladenosine sodium salt Sigma Aldrich # M2780 ethanol precipitated and resuspended with RNase free water. The eluted RNA and input RNA were subjected to 3’pre adenylated DNA liker ligation with T4 RNA ligase2 truncated KQ NEB #M0373L at 16 oC over night. The sequencing library was constructed with bromodeoxyuridine BrdU CLIP protocol described in Weyn Vanhentenryck et al. 2014 with improved RT primers indicated in Ke et al. 2015. Embryos from the AB wild type strain were obtained from the NMBU zebrafish facility where the zebrafish were kept at 28±1°C on a 14 10 hour light dark cycle at a density of 5 10 fish/L. System water SW was prepared from particle and active charcoal filtrated tap water deionized by reverse osmosis RO and kept sterile by UV irradiation. The water was conditioned to a conductivity of 500µS/cm general hardness GH 4 5 and pH 7.5 by addition of 155g synthetic sea salt Instant Ocean Blacksburg USA 53g sodium carbonate and 15g calcium chloride Sigma Aldrich per liter RO water. Adult fish were fed with Gemma Micro 300 Skretting Stavanger Norway dry feed twice a day and live artemia Scanbur Karlslunde Denmark once a day. Health monitoring was by daily inspection use of sentinel fish sent for pathology ZIRC Eugene Oregon and water microbiology analysis NMBU Vetbio Oslo every six month. Adult fish were allowed to mate for 30 minutes in standard 1L breeding tanks Aquatic Habitats Apopka FL. Harvested embryos were kept in autoclaved SW at 28 °C harvested by snap freezing liquid nitrogen and visually controlled for stage and lack of abnormalities at the selected time points. All experiments were performed according to Norwegian Animal Welfare Act 2009 the EU Directive 2010/63. developmental stage:Shield stage embryos|strain:AB wild type GSM2390028 GSM2390028: input shield rep2; Danio rerio; RIP Seq GSM2390028   1 We isolated total RNA from 4 stages 1 cell/20 minutes post fertilization 2 4 and 6 hpf in batches of 200 embryos using TRIzol Invitrogen cat.no. 15596 018. We added ERCC spike in RNA TermoFisher scientific cat.no. 4456740 to the trizol. For each sequencing library two batches of total RNA from 200 embryos were merged and enriched for polyA+ RNA using Dynabeads® mRNA Purification Kit Ambion #61006. The m6A RIP experiment was carried out as previously described Ke et al. 2015 and the protocol can be found in supplementary file 1. Briefly polyA+ enriched RNA was partially fragmented by alkaline hydrolysis ethanol precipitated and SDS PAGE size selected for 20 80nt. Part of the fragmented RNA was used as input and the rest was immunoprecipitated at 4oC for 2 h using Dynabeads Protein A Life Technologies # 10008D conjugated anti m6A antibody Synaptic systems # 202003. post stringent washing the bound RNA was eluted with 0.5mg/ml N6 methyladenosine sodium salt Sigma Aldrich # M2780 ethanol precipitated and resuspended with RNase free water. The eluted RNA and input RNA were subjected to 3’pre adenylated DNA liker ligation with T4 RNA ligase2 truncated KQ NEB #M0373L at 16 oC over night. The sequencing library was constructed with bromodeoxyuridine BrdU CLIP protocol described in Weyn Vanhentenryck et al. 2014 with improved RT primers indicated in Ke et al. 2015. GEO Accession:GSM2390028 RIP-Seq TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2000   SRP093295     input_shield_rep2.fastq.gz fastq 6559838034.0 69785511.0 GSM2390028 r1 0:94 A:1785772532;C:1552902091;G:1571407685;T:1649464013;N:291713 94       1785772532 1552902091 1571407685 1649464013 291713 SRX2345570 SRS1796136 SRA492943 GEO Klungland Lab, Dept of microbiology, Oslo University Hospital 1 0.03627   0.01   0.96676   0.66777   94   B   usable mapping rate illumina hiseq_era unknown poly_a unknown bulk unknown unknown   Norway 2016-11-14 Gastrula Embryo Embryo Imprecise All anatomical structures
41549 41549 SRR5017074 SRX2345569 SRS1796134 SRP093295 PRJNA353372 N6 methyladenosine dynamics during early vertebrate embryogenesis GSE89815 Other Early vertebrate embryogenesis is characterized by extensive post transcriptional regulation during the maternal to zygotic transition. The N6 methyladenosine m6A modifications on mRNA has been shown to affect both translation and stability of transcripts. Here we investigate the m6A topology during early vertebrate embryogenesis and its association with RNA stability translation efficiency and effect on miR 430 degradation kinetics. Notably we find a strong association of m6A with cytoplasmic polyadenylation and translational efficiency prior to zygotic genome activation. Genes required for zygotic genome activation such as nanog and pou5f3 display dynamic m6A levels. post zygotic genome activation m6A is associated with improved stability and dampens the effect of miR 430 mediated degradation. Through sequence analyses we identified enrichment of motifs for RNA binding proteins involved in translational regulation and RNA degradation. We propose a role for m6A in multiple mRNA regulatory mechanisms for the first time in an in vivo system and improve our understanding of the combinatorial code behind the complex post transcriptional regulation of reprogramming during early vertebrate development. Overall design: Examination of m6A in four different developmental stages       input shield rep1 GSM2390027   tissue:Embryos|developmental stage:Shield stage embryos|strain:AB wild type input shield rep1 Base calling The sequencing output from each of the 4 lanes the sequencing run was trimmed 7 nucleotides in the 5’end and de multiplexed 4 samples in each lane. Reads were mapped to Zv10 using the STAR aligner Dobin et al. 2013 with options seedSearchStartLmax 15 clip3pNbases 10 clip5pNbases 10 outFilterMultimapNmax 20 outFilterMismatchNoverLmax 0.05 outFilterMatchNminOverLread 0.0 outFilterMatchNmin 15 outFilterScoreMinOverLread 0.0. We performed peak calling and detection of differentially methylated genes using ExomePeak Meng et al. 2013. Genome build: Zv10 Supplementary files format and content: Bed files with enriched regions from each developmental stage and txt file with raw and normalized read counts for each gene using the input samples Embryos   We isolated total RNA from 4 stages 1 cell/20 minutes post fertilization 2 4 and 6 hpf in batches of 200 embryos using TRIzol Invitrogen cat.no. 15596 018. We added ERCC spike in RNA TermoFisher scientific cat.no. 4456740 to the trizol. For each sequencing library two batches of total RNA from 200 embryos were merged and enriched for polyA+ RNA using Dynabeads® mRNA Purification Kit Ambion #61006. The m6A RIP experiment was carried out as previously described Ke et al. 2015 and the protocol can be found in supplementary file 1. Briefly polyA+ enriched RNA was partially fragmented by alkaline hydrolysis ethanol precipitated and SDS PAGE size selected for 20 80nt. Part of the fragmented RNA was used as input and the rest was immunoprecipitated at 4oC for 2 h using Dynabeads Protein A Life Technologies # 10008D conjugated anti m6A antibody Synaptic systems # 202003. post stringent washing the bound RNA was eluted with 0.5mg/ml N6 methyladenosine sodium salt Sigma Aldrich # M2780 ethanol precipitated and resuspended with RNase free water. The eluted RNA and input RNA were subjected to 3’pre adenylated DNA liker ligation with T4 RNA ligase2 truncated KQ NEB #M0373L at 16 oC over night. The sequencing library was constructed with bromodeoxyuridine BrdU CLIP protocol described in Weyn Vanhentenryck et al. 2014 with improved RT primers indicated in Ke et al. 2015. Embryos from the AB wild type strain were obtained from the NMBU zebrafish facility where the zebrafish were kept at 28±1°C on a 14 10 hour light dark cycle at a density of 5 10 fish/L. System water SW was prepared from particle and active charcoal filtrated tap water deionized by reverse osmosis RO and kept sterile by UV irradiation. The water was conditioned to a conductivity of 500µS/cm general hardness GH 4 5 and pH 7.5 by addition of 155g synthetic sea salt Instant Ocean Blacksburg USA 53g sodium carbonate and 15g calcium chloride Sigma Aldrich per liter RO water. Adult fish were fed with Gemma Micro 300 Skretting Stavanger Norway dry feed twice a day and live artemia Scanbur Karlslunde Denmark once a day. Health monitoring was by daily inspection use of sentinel fish sent for pathology ZIRC Eugene Oregon and water microbiology analysis NMBU Vetbio Oslo every six month. Adult fish were allowed to mate for 30 minutes in standard 1L breeding tanks Aquatic Habitats Apopka FL. Harvested embryos were kept in autoclaved SW at 28 °C harvested by snap freezing liquid nitrogen and visually controlled for stage and lack of abnormalities at the selected time points. All experiments were performed according to Norwegian Animal Welfare Act 2009 the EU Directive 2010/63. developmental stage:Shield stage embryos|strain:AB wild type GSM2390027 GSM2390027: input shield rep1; Danio rerio; RIP Seq GSM2390027   1 We isolated total RNA from 4 stages 1 cell/20 minutes post fertilization 2 4 and 6 hpf in batches of 200 embryos using TRIzol Invitrogen cat.no. 15596 018. We added ERCC spike in RNA TermoFisher scientific cat.no. 4456740 to the trizol. For each sequencing library two batches of total RNA from 200 embryos were merged and enriched for polyA+ RNA using Dynabeads® mRNA Purification Kit Ambion #61006. The m6A RIP experiment was carried out as previously described Ke et al. 2015 and the protocol can be found in supplementary file 1. Briefly polyA+ enriched RNA was partially fragmented by alkaline hydrolysis ethanol precipitated and SDS PAGE size selected for 20 80nt. Part of the fragmented RNA was used as input and the rest was immunoprecipitated at 4oC for 2 h using Dynabeads Protein A Life Technologies # 10008D conjugated anti m6A antibody Synaptic systems # 202003. post stringent washing the bound RNA was eluted with 0.5mg/ml N6 methyladenosine sodium salt Sigma Aldrich # M2780 ethanol precipitated and resuspended with RNase free water. The eluted RNA and input RNA were subjected to 3’pre adenylated DNA liker ligation with T4 RNA ligase2 truncated KQ NEB #M0373L at 16 oC over night. The sequencing library was constructed with bromodeoxyuridine BrdU CLIP protocol described in Weyn Vanhentenryck et al. 2014 with improved RT primers indicated in Ke et al. 2015. GEO Accession:GSM2390027 RIP-Seq TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2000   SRP093295     input_shield_rep1.fastq.gz fastq 3765873214.0 40062481.0 GSM2390027 r1 0:94 A:1002751444;C:902509483;G:908627718;T:951814220;N:170349 94       1002751444 902509483 908627718 951814220 170349 SRX2345569 SRS1796134 SRA492943 GEO Klungland Lab, Dept of microbiology, Oslo University Hospital 1 0.03936   0.01095   0.96518   0.68725   94   B   usable mapping rate illumina hiseq_era unknown poly_a unknown bulk unknown unknown   Norway 2016-11-14 Gastrula Embryo Embryo Imprecise All anatomical structures
41550 41550 SRR5017073 SRX2345568 SRS1796132 SRP093295 PRJNA353372 N6 methyladenosine dynamics during early vertebrate embryogenesis GSE89815 Other Early vertebrate embryogenesis is characterized by extensive post transcriptional regulation during the maternal to zygotic transition. The N6 methyladenosine m6A modifications on mRNA has been shown to affect both translation and stability of transcripts. Here we investigate the m6A topology during early vertebrate embryogenesis and its association with RNA stability translation efficiency and effect on miR 430 degradation kinetics. Notably we find a strong association of m6A with cytoplasmic polyadenylation and translational efficiency prior to zygotic genome activation. Genes required for zygotic genome activation such as nanog and pou5f3 display dynamic m6A levels. post zygotic genome activation m6A is associated with improved stability and dampens the effect of miR 430 mediated degradation. Through sequence analyses we identified enrichment of motifs for RNA binding proteins involved in translational regulation and RNA degradation. We propose a role for m6A in multiple mRNA regulatory mechanisms for the first time in an in vivo system and improve our understanding of the combinatorial code behind the complex post transcriptional regulation of reprogramming during early vertebrate development. Overall design: Examination of m6A in four different developmental stages       ip shield rep2 GSM2390026   tissue:Embryos|developmental stage:Shield stage embryos|strain:AB wild type ip shield rep2 Base calling The sequencing output from each of the 4 lanes the sequencing run was trimmed 7 nucleotides in the 5’end and de multiplexed 4 samples in each lane. Reads were mapped to Zv10 using the STAR aligner Dobin et al. 2013 with options seedSearchStartLmax 15 clip3pNbases 10 clip5pNbases 10 outFilterMultimapNmax 20 outFilterMismatchNoverLmax 0.05 outFilterMatchNminOverLread 0.0 outFilterMatchNmin 15 outFilterScoreMinOverLread 0.0. We performed peak calling and detection of differentially methylated genes using ExomePeak Meng et al. 2013. Genome build: Zv10 Supplementary files format and content: Bed files with enriched regions from each developmental stage and txt file with raw and normalized read counts for each gene using the input samples Embryos   We isolated total RNA from 4 stages 1 cell/20 minutes post fertilization 2 4 and 6 hpf in batches of 200 embryos using TRIzol Invitrogen cat.no. 15596 018. We added ERCC spike in RNA TermoFisher scientific cat.no. 4456740 to the trizol. For each sequencing library two batches of total RNA from 200 embryos were merged and enriched for polyA+ RNA using Dynabeads® mRNA Purification Kit Ambion #61006. The m6A RIP experiment was carried out as previously described Ke et al. 2015 and the protocol can be found in supplementary file 1. Briefly polyA+ enriched RNA was partially fragmented by alkaline hydrolysis ethanol precipitated and SDS PAGE size selected for 20 80nt. Part of the fragmented RNA was used as input and the rest was immunoprecipitated at 4oC for 2 h using Dynabeads Protein A Life Technologies # 10008D conjugated anti m6A antibody Synaptic systems # 202003. post stringent washing the bound RNA was eluted with 0.5mg/ml N6 methyladenosine sodium salt Sigma Aldrich # M2780 ethanol precipitated and resuspended with RNase free water. The eluted RNA and input RNA were subjected to 3’pre adenylated DNA liker ligation with T4 RNA ligase2 truncated KQ NEB #M0373L at 16 oC over night. The sequencing library was constructed with bromodeoxyuridine BrdU CLIP protocol described in Weyn Vanhentenryck et al. 2014 with improved RT primers indicated in Ke et al. 2015. Embryos from the AB wild type strain were obtained from the NMBU zebrafish facility where the zebrafish were kept at 28±1°C on a 14 10 hour light dark cycle at a density of 5 10 fish/L. System water SW was prepared from particle and active charcoal filtrated tap water deionized by reverse osmosis RO and kept sterile by UV irradiation. The water was conditioned to a conductivity of 500µS/cm general hardness GH 4 5 and pH 7.5 by addition of 155g synthetic sea salt Instant Ocean Blacksburg USA 53g sodium carbonate and 15g calcium chloride Sigma Aldrich per liter RO water. Adult fish were fed with Gemma Micro 300 Skretting Stavanger Norway dry feed twice a day and live artemia Scanbur Karlslunde Denmark once a day. Health monitoring was by daily inspection use of sentinel fish sent for pathology ZIRC Eugene Oregon and water microbiology analysis NMBU Vetbio Oslo every six month. Adult fish were allowed to mate for 30 minutes in standard 1L breeding tanks Aquatic Habitats Apopka FL. Harvested embryos were kept in autoclaved SW at 28 °C harvested by snap freezing liquid nitrogen and visually controlled for stage and lack of abnormalities at the selected time points. All experiments were performed according to Norwegian Animal Welfare Act 2009 the EU Directive 2010/63. developmental stage:Shield stage embryos|strain:AB wild type GSM2390026 GSM2390026: ip shield rep2; Danio rerio; RIP Seq GSM2390026   1 We isolated total RNA from 4 stages 1 cell/20 minutes post fertilization 2 4 and 6 hpf in batches of 200 embryos using TRIzol Invitrogen cat.no. 15596 018. We added ERCC spike in RNA TermoFisher scientific cat.no. 4456740 to the trizol. For each sequencing library two batches of total RNA from 200 embryos were merged and enriched for polyA+ RNA using Dynabeads® mRNA Purification Kit Ambion #61006. The m6A RIP experiment was carried out as previously described Ke et al. 2015 and the protocol can be found in supplementary file 1. Briefly polyA+ enriched RNA was partially fragmented by alkaline hydrolysis ethanol precipitated and SDS PAGE size selected for 20 80nt. Part of the fragmented RNA was used as input and the rest was immunoprecipitated at 4oC for 2 h using Dynabeads Protein A Life Technologies # 10008D conjugated anti m6A antibody Synaptic systems # 202003. post stringent washing the bound RNA was eluted with 0.5mg/ml N6 methyladenosine sodium salt Sigma Aldrich # M2780 ethanol precipitated and resuspended with RNase free water. The eluted RNA and input RNA were subjected to 3’pre adenylated DNA liker ligation with T4 RNA ligase2 truncated KQ NEB #M0373L at 16 oC over night. The sequencing library was constructed with bromodeoxyuridine BrdU CLIP protocol described in Weyn Vanhentenryck et al. 2014 with improved RT primers indicated in Ke et al. 2015. GEO Accession:GSM2390026 RIP-Seq TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2000   SRP093295     ip_shield_rep2.fastq fastq 3855863926.0 41019829.0 GSM2390026 r1 0:94 A:1044010018;C:931492803;G:952890289;T:926349254;N:1121562 94       1044010018 931492803 952890289 926349254 1121562 SRX2345568 SRS1796132 SRA492943 GEO Klungland Lab, Dept of microbiology, Oslo University Hospital 1 0.03311   0.00481   0.96337   0.68612   94   B   usable mapping rate illumina hiseq_era unknown poly_a unknown bulk unknown unknown   Norway 2016-11-14 Gastrula Embryo Embryo Imprecise All anatomical structures
41551 41551 SRR5017072 SRX2345567 SRS1796131 SRP093295 PRJNA353372 N6 methyladenosine dynamics during early vertebrate embryogenesis GSE89815 Other Early vertebrate embryogenesis is characterized by extensive post transcriptional regulation during the maternal to zygotic transition. The N6 methyladenosine m6A modifications on mRNA has been shown to affect both translation and stability of transcripts. Here we investigate the m6A topology during early vertebrate embryogenesis and its association with RNA stability translation efficiency and effect on miR 430 degradation kinetics. Notably we find a strong association of m6A with cytoplasmic polyadenylation and translational efficiency prior to zygotic genome activation. Genes required for zygotic genome activation such as nanog and pou5f3 display dynamic m6A levels. post zygotic genome activation m6A is associated with improved stability and dampens the effect of miR 430 mediated degradation. Through sequence analyses we identified enrichment of motifs for RNA binding proteins involved in translational regulation and RNA degradation. We propose a role for m6A in multiple mRNA regulatory mechanisms for the first time in an in vivo system and improve our understanding of the combinatorial code behind the complex post transcriptional regulation of reprogramming during early vertebrate development. Overall design: Examination of m6A in four different developmental stages       ip shield rep1 GSM2390025   tissue:Embryos|developmental stage:Shield stage embryos|strain:AB wild type ip shield rep1 Base calling The sequencing output from each of the 4 lanes the sequencing run was trimmed 7 nucleotides in the 5’end and de multiplexed 4 samples in each lane. Reads were mapped to Zv10 using the STAR aligner Dobin et al. 2013 with options seedSearchStartLmax 15 clip3pNbases 10 clip5pNbases 10 outFilterMultimapNmax 20 outFilterMismatchNoverLmax 0.05 outFilterMatchNminOverLread 0.0 outFilterMatchNmin 15 outFilterScoreMinOverLread 0.0. We performed peak calling and detection of differentially methylated genes using ExomePeak Meng et al. 2013. Genome build: Zv10 Supplementary files format and content: Bed files with enriched regions from each developmental stage and txt file with raw and normalized read counts for each gene using the input samples Embryos   We isolated total RNA from 4 stages 1 cell/20 minutes post fertilization 2 4 and 6 hpf in batches of 200 embryos using TRIzol Invitrogen cat.no. 15596 018. We added ERCC spike in RNA TermoFisher scientific cat.no. 4456740 to the trizol. For each sequencing library two batches of total RNA from 200 embryos were merged and enriched for polyA+ RNA using Dynabeads® mRNA Purification Kit Ambion #61006. The m6A RIP experiment was carried out as previously described Ke et al. 2015 and the protocol can be found in supplementary file 1. Briefly polyA+ enriched RNA was partially fragmented by alkaline hydrolysis ethanol precipitated and SDS PAGE size selected for 20 80nt. Part of the fragmented RNA was used as input and the rest was immunoprecipitated at 4oC for 2 h using Dynabeads Protein A Life Technologies # 10008D conjugated anti m6A antibody Synaptic systems # 202003. post stringent washing the bound RNA was eluted with 0.5mg/ml N6 methyladenosine sodium salt Sigma Aldrich # M2780 ethanol precipitated and resuspended with RNase free water. The eluted RNA and input RNA were subjected to 3’pre adenylated DNA liker ligation with T4 RNA ligase2 truncated KQ NEB #M0373L at 16 oC over night. The sequencing library was constructed with bromodeoxyuridine BrdU CLIP protocol described in Weyn Vanhentenryck et al. 2014 with improved RT primers indicated in Ke et al. 2015. Embryos from the AB wild type strain were obtained from the NMBU zebrafish facility where the zebrafish were kept at 28±1°C on a 14 10 hour light dark cycle at a density of 5 10 fish/L. System water SW was prepared from particle and active charcoal filtrated tap water deionized by reverse osmosis RO and kept sterile by UV irradiation. The water was conditioned to a conductivity of 500µS/cm general hardness GH 4 5 and pH 7.5 by addition of 155g synthetic sea salt Instant Ocean Blacksburg USA 53g sodium carbonate and 15g calcium chloride Sigma Aldrich per liter RO water. Adult fish were fed with Gemma Micro 300 Skretting Stavanger Norway dry feed twice a day and live artemia Scanbur Karlslunde Denmark once a day. Health monitoring was by daily inspection use of sentinel fish sent for pathology ZIRC Eugene Oregon and water microbiology analysis NMBU Vetbio Oslo every six month. Adult fish were allowed to mate for 30 minutes in standard 1L breeding tanks Aquatic Habitats Apopka FL. Harvested embryos were kept in autoclaved SW at 28 °C harvested by snap freezing liquid nitrogen and visually controlled for stage and lack of abnormalities at the selected time points. All experiments were performed according to Norwegian Animal Welfare Act 2009 the EU Directive 2010/63. developmental stage:Shield stage embryos|strain:AB wild type GSM2390025 GSM2390025: ip shield rep1; Danio rerio; RIP Seq GSM2390025   1 We isolated total RNA from 4 stages 1 cell/20 minutes post fertilization 2 4 and 6 hpf in batches of 200 embryos using TRIzol Invitrogen cat.no. 15596 018. We added ERCC spike in RNA TermoFisher scientific cat.no. 4456740 to the trizol. For each sequencing library two batches of total RNA from 200 embryos were merged and enriched for polyA+ RNA using Dynabeads® mRNA Purification Kit Ambion #61006. The m6A RIP experiment was carried out as previously described Ke et al. 2015 and the protocol can be found in supplementary file 1. Briefly polyA+ enriched RNA was partially fragmented by alkaline hydrolysis ethanol precipitated and SDS PAGE size selected for 20 80nt. Part of the fragmented RNA was used as input and the rest was immunoprecipitated at 4oC for 2 h using Dynabeads Protein A Life Technologies # 10008D conjugated anti m6A antibody Synaptic systems # 202003. post stringent washing the bound RNA was eluted with 0.5mg/ml N6 methyladenosine sodium salt Sigma Aldrich # M2780 ethanol precipitated and resuspended with RNase free water. The eluted RNA and input RNA were subjected to 3’pre adenylated DNA liker ligation with T4 RNA ligase2 truncated KQ NEB #M0373L at 16 oC over night. The sequencing library was constructed with bromodeoxyuridine BrdU CLIP protocol described in Weyn Vanhentenryck et al. 2014 with improved RT primers indicated in Ke et al. 2015. GEO Accession:GSM2390025 RIP-Seq TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2000   SRP093295     ip_shield_rep1.fastq.gz fastq 4107624502.0 43698133.0 GSM2390025 r1 0:94 A:1087077899;C:996101257;G:1020588721;T:1002667943;N:1188682 94       1087077899 996101257 1020588721 1002667943 1188682 SRX2345567 SRS1796131 SRA492943 GEO Klungland Lab, Dept of microbiology, Oslo University Hospital 1 0.01898   0.00264   0.97238   0.59809   94   B   usable mapping rate illumina hiseq_era unknown poly_a unknown bulk unknown unknown   Norway 2016-11-14 Gastrula Embryo Embryo Imprecise All anatomical structures
44644 44644 SRR6268204 SRX3374372 SRS2671596 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   oocyte A+ 7h GSM2845358   source name:zebrafish oocyte|developmental stage:7h|tissue:oocyte oocyte A+ 7h Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish oocyte Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:7h|tissue:oocyte GSM2845358 GSM2845358: oocyte A+ 7h; Danio rerio; OTHER GSM2845358   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845358 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina MiSeq   SRP124609     AR22-SL21.fastq.gz fastq 463495840.0 2896849.0 GSM2845358 r1 0:160 1:0 A:145685088;C:104925443;G:89232572;T:123650704;N:2033 160 0     145685088 104925443 89232572 123650704 2033 SRX3374372 SRS2671596 SRA629220 GEO Broad Institute 1 4e-05   1e-05   0.99991   0.5   160   T   under 1.2% mapping rate illumina miseq unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Oocyte Reproductive System
44647 44647 SRR6268201 SRX3374369 SRS2671601 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   oocyte A 7h GSM2845355   source name:zebrafish oocyte|developmental stage:7h|tissue:oocyte oocyte A 7h Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish oocyte Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:7h|tissue:oocyte GSM2845355 GSM2845355: oocyte A 7h; Danio rerio; OTHER GSM2845355   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845355 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina MiSeq   SRP124609     AR19-SL40.fastq.gz fastq 437084640.0 2731779.0 GSM2845355 r1 0:160 1:0 A:138575507;C:98512376;G:80808399;T:119186452;N:1906 160 0     138575507 98512376 80808399 119186452 1906 SRX3374369 SRS2671601 SRA629220 GEO Broad Institute 1 2e-05   0.0   0.99995   1.0   160   T   under 1.2% mapping rate illumina miseq unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Oocyte Reproductive System
44650 44650 SRR6268198 SRX3374366 SRS2671592 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   rep A 10h GSM2845352   source name:zebrafish embryos|developmental stage:10hpf|tissue:embryo rep A 10h Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:10hpf|tissue:embryo GSM2845352 GSM2845352: rep A 10h; Danio rerio; OTHER GSM2845352   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845352 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina MiSeq   SRP124609     10hpf.fastq.gz fastq 267594560.0 1672466.0 GSM2845352 r1 0:160 1:0 A:87249429;C:58556737;G:48918401;T:72845229;N:24764 160 0     87249429 58556737 48918401 72845229 24764 SRX3374366 SRS2671592 SRA629220 GEO Broad Institute 1 2e-05   0.0   0.99995   0.5   160   T   under 1.2% mapping rate illumina miseq unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44651 44651 SRR6268197 SRX3374365 SRS2671591 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   rep A 8h GSM2845351   source name:zebrafish embryos|developmental stage:8hpf|tissue:embryo rep A 8h Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:8hpf|tissue:embryo GSM2845351 GSM2845351: rep A 8h; Danio rerio; OTHER GSM2845351   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845351 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina MiSeq   SRP124609     8hpf.fastq.gz fastq 277648000.0 1735300.0 GSM2845351 r1 0:160 1:0 A:88447999;C:60766591;G:51931752;T:76474911;N:26747 160 0     88447999 60766591 51931752 76474911 26747 SRX3374365 SRS2671591 SRA629220 GEO Broad Institute 1 0.0   0.0   1.0       160   T   under 1.2% mapping rate illumina miseq unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44652 44652 SRR6268196 SRX3374364 SRS2671590 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   rep A 6h.3 GSM2845350   source name:zebrafish embryos|developmental stage:6hpf|tissue:embryo rep A 6h.3 Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:6hpf|tissue:embryo GSM2845350 GSM2845350: rep A 6h.3; Danio rerio; OTHER GSM2845350   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845350 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina MiSeq   SRP124609     6hpf.fastq.gz fastq 321713920.0 2010712.0 GSM2845350 r1 0:160 1:0 A:108106130;C:67673948;G:57401886;T:88504064;N:27892 160 0     108106130 67673948 57401886 88504064 27892 SRX3374364 SRS2671590 SRA629220 GEO Broad Institute 1 4e-05   0.0   0.99985   0.57142   160   T   under 1.2% mapping rate illumina miseq unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44653 44653 SRR6268195 SRX3374363 SRS2671589 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   rep A 6h.2 GSM2845349   source name:zebrafish embryos|developmental stage:6hpf|tissue:embryo rep A 6h.2 Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:6hpf|tissue:embryo GSM2845349 GSM2845349: rep A 6h.2; Danio rerio; OTHER GSM2845349   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845349 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina MiSeq   SRP124609     4hpf-3.fastq.gz fastq 469592000.0 2934950.0 GSM2845349 r1 0:160 1:0 A:149657114;C:104627528;G:87984247;T:127285382;N:37729 160 0     149657114 104627528 87984247 127285382 37729 SRX3374363 SRS2671589 SRA629220 GEO Broad Institute 1 1e-05   0.0   0.99997   0.0   160   T   under 1.2% mapping rate illumina miseq unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44654 44654 SRR6268194 SRX3374362 SRS2671588 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   rep A 6h.1 GSM2845348   source name:zebrafish embryos|developmental stage:6hpf|tissue:embryo rep A 6h.1 Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:6hpf|tissue:embryo GSM2845348 GSM2845348: rep A 6h.1; Danio rerio; OTHER GSM2845348   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845348 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina MiSeq   SRP124609     4hpf-2.fastq.gz fastq 450388640.0 2814929.0 GSM2845348 r1 0:160 1:0 A:143486047;C:99354304;G:85285329;T:122222711;N:40249 160 0     143486047 99354304 85285329 122222711 40249 SRX3374362 SRS2671588 SRA629220 GEO Broad Institute 1 3e-05   0.0   0.99991   0.5   160   T   under 1.2% mapping rate illumina miseq unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44660 44660 SRR6268188 SRX3374356 SRS2671583 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   techrep A+ 10h.2 GSM2845342   source name:zebrafish embryos|developmental stage:10hpf|tissue:embryo techrep A+ 10h.2 Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:10hpf|tissue:embryo GSM2845342 GSM2845342: techrep A+ 10h.2; Danio rerio; OTHER GSM2845342   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845342 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina MiSeq   SRP124609     AR24_S24.fastq.gz fastq 152993736.0 910677.0 GSM2845342 r1 0:168 1:0 A:51571958;C:32339728;G:27945885;T:41135710;N:455 168 0     51571958 32339728 27945885 41135710 455 SRX3374356 SRS2671583 SRA629220 GEO Broad Institute 1 5e-05   0.0   0.99989   0.5   168   T   under 1.2% mapping rate illumina miseq unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44661 44661 SRR6268187 SRX3374355 SRS2671580 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   techrep A+ 9h.2 GSM2845341   source name:zebrafish embryos|developmental stage:9hpf|tissue:embryo techrep A+ 9h.2 Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:9hpf|tissue:embryo GSM2845341 GSM2845341: techrep A+ 9h.2; Danio rerio; OTHER GSM2845341   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845341 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina MiSeq   SRP124609     AR23_S23.fastq.gz fastq 170881704.0 1017153.0 GSM2845341 r1 0:168 1:0 A:58715337;C:35449423;G:30006312;T:46710126;N:506 168 0     58715337 35449423 30006312 46710126 506 SRX3374355 SRS2671580 SRA629220 GEO Broad Institute 1 2e-05   0.0   0.99995   0.5   168   T   under 1.2% mapping rate illumina miseq unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44662 44662 SRR6268186 SRX3374354 SRS2671582 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   techrep A+ 8h.2 GSM2845340   source name:zebrafish embryos|developmental stage:8hpf|tissue:embryo techrep A+ 8h.2 Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:8hpf|tissue:embryo GSM2845340 GSM2845340: techrep A+ 8h.2; Danio rerio; OTHER GSM2845340   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845340 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina MiSeq   SRP124609     AR22_S22.fastq.gz fastq 162500520.0 967265.0 GSM2845340 r1 0:168 1:0 A:56281233;C:33095421;G:28617222;T:44506190;N:454 168 0     56281233 33095421 28617222 44506190 454 SRX3374354 SRS2671582 SRA629220 GEO Broad Institute 1 2e-05   0.0   0.99995   1.0   168   T   under 1.2% mapping rate illumina miseq unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44663 44663 SRR6268185 SRX3374353 SRS2671581 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   techrep A+ 7h.2 GSM2845339   source name:zebrafish embryos|developmental stage:7hpf|tissue:embryo techrep A+ 7h.2 Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:7hpf|tissue:embryo GSM2845339 GSM2845339: techrep A+ 7h.2; Danio rerio; OTHER GSM2845339   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845339 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina MiSeq   SRP124609     AR21_S21.fastq.gz fastq 177185400.0 1054675.0 GSM2845339 r1 0:168 1:0 A:61487643;C:35736507;G:31141191;T:48819498;N:561 168 0     61487643 35736507 31141191 48819498 561 SRX3374353 SRS2671581 SRA629220 GEO Broad Institute 1 3e-05   0.0   0.99995   1.0   168   T   under 1.2% mapping rate illumina miseq unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44664 44664 SRR6268184 SRX3374352 SRS2671579 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   techrep A+ 6h.2.2 GSM2845338   source name:zebrafish embryos|developmental stage:6hpf|tissue:embryo techrep A+ 6h.2.2 Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:6hpf|tissue:embryo GSM2845338 GSM2845338: techrep A+ 6h.2.2; Danio rerio; OTHER GSM2845338   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845338 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina MiSeq   SRP124609     AR20_S20.fastq.gz fastq 182756784.0 1087838.0 GSM2845338 r1 0:168 1:0 A:60314822;C:39299119;G:34916612;T:48225663;N:568 168 0     60314822 39299119 34916612 48225663 568 SRX3374352 SRS2671579 SRA629220 GEO Broad Institute 1 2e-05   0.0   0.99997   0.0   168   T   under 1.2% mapping rate illumina miseq unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44665 44665 SRR6268183 SRX3374351 SRS2671599 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   techrep A+ 6h.2.1 GSM2845337   source name:zebrafish embryos|developmental stage:6hpf|tissue:embryo techrep A+ 6h.2.1 Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:6hpf|tissue:embryo GSM2845337 GSM2845337: techrep A+ 6h.2.1; Danio rerio; OTHER GSM2845337   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845337 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina MiSeq   SRP124609     AR19_S19.fastq.gz fastq 160068720.0 952790.0 GSM2845337 r1 0:168 1:0 A:55382285;C:32355561;G:29019311;T:43311149;N:414 168 0     55382285 32355561 29019311 43311149 414 SRX3374351 SRS2671599 SRA629220 GEO Broad Institute 1 2e-05   0.0   0.99995   1.0   168   T   under 1.2% mapping rate illumina miseq unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44672 44672 SRR6268176 SRX3374344 SRS2671571 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   techrep A+ 10h.1 GSM2845330   source name:zebrafish embryos|developmental stage:10hpf|tissue:embryo techrep A+ 10h.1 Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:10hpf|tissue:embryo GSM2845330 GSM2845330: techrep A+ 10h.1; Danio rerio; OTHER GSM2845330   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845330 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina MiSeq   SRP124609     AR12_S12.fastq.gz fastq 183198288.0 1090466.0 GSM2845330 r1 0:168 1:0 A:62131603;C:37866401;G:33149583;T:50050141;N:560 168 0     62131603 37866401 33149583 50050141 560 SRX3374344 SRS2671571 SRA629220 GEO Broad Institute 1 0.0   0.0   1.0       168   T   under 1.2% mapping rate illumina miseq unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44673 44673 SRR6268175 SRX3374343 SRS2671572 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   techrep A+ 9h.1 GSM2845329   source name:zebrafish embryos|developmental stage:9hpf|tissue:embryo techrep A+ 9h.1 Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:9hpf|tissue:embryo GSM2845329 GSM2845329: techrep A+ 9h.1; Danio rerio; OTHER GSM2845329   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845329 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina MiSeq   SRP124609     AR11_S11.fastq.gz fastq 254056656.0 1512242.0 GSM2845329 r1 0:168 1:0 A:85470904;C:52657639;G:46610564;T:69316784;N:765 168 0     85470904 52657639 46610564 69316784 765 SRX3374343 SRS2671572 SRA629220 GEO Broad Institute 1 2e-05   0.0   0.99997   1.0   168   T   under 1.2% mapping rate illumina miseq unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44674 44674 SRR6268174 SRX3374342 SRS2671570 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   techrep A+ 8h.1 GSM2845328   source name:zebrafish embryos|developmental stage:8hpf|tissue:embryo techrep A+ 8h.1 Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:8hpf|tissue:embryo GSM2845328 GSM2845328: techrep A+ 8h.1; Danio rerio; OTHER GSM2845328   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845328 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina MiSeq   SRP124609     AR10_S10.fastq.gz fastq 219042936.0 1303827.0 GSM2845328 r1 0:168 1:0 A:72968024;C:46155718;G:40847360;T:59071152;N:682 168 0     72968024 46155718 40847360 59071152 682 SRX3374342 SRS2671570 SRA629220 GEO Broad Institute 1 0.0   0.0   1.0       168   T   under 1.2% mapping rate illumina miseq unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44675 44675 SRR6268173 SRX3374341 SRS2671569 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   techrep A+ 7h.1 GSM2845327   source name:zebrafish embryos|developmental stage:7hpf|tissue:embryo techrep A+ 7h.1 Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:7hpf|tissue:embryo GSM2845327 GSM2845327: techrep A+ 7h.1; Danio rerio; OTHER GSM2845327   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845327 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina MiSeq   SRP124609     AR09_S9.fastq.gz fastq 295385160.0 1758245.0 GSM2845327 r1 0:168 1:0 A:96898707;C:64356190;G:55256265;T:78873123;N:875 168 0     96898707 64356190 55256265 78873123 875 SRX3374341 SRS2671569 SRA629220 GEO Broad Institute 1 1e-05   0.0   0.99997   0.0   168   T   under 1.2% mapping rate illumina miseq unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44676 44676 SRR6268172 SRX3374340 SRS2671568 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   techrep A+ 6h.1.2 GSM2845326   source name:zebrafish embryos|developmental stage:6hpf|tissue:embryo techrep A+ 6h.1.2 Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:6hpf|tissue:embryo GSM2845326 GSM2845326: techrep A+ 6h.1.2; Danio rerio; OTHER GSM2845326   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845326 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina MiSeq   SRP124609     AR08_S8.fastq.gz fastq 200314464.0 1192348.0 GSM2845326 r1 0:168 1:0 A:69255469;C:40029074;G:36656767;T:54372650;N:504 168 0     69255469 40029074 36656767 54372650 504 SRX3374340 SRS2671568 SRA629220 GEO Broad Institute 1 0.0   0.0   1.0       168   T   under 1.2% mapping rate illumina miseq unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44677 44677 SRR6268171 SRX3374339 SRS2671567 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   techrep A+ 6h.1.1 GSM2845325   source name:zebrafish embryos|developmental stage:6hpf|tissue:embryo techrep A+ 6h.1.1 Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:6hpf|tissue:embryo GSM2845325 GSM2845325: techrep A+ 6h.1.1; Danio rerio; OTHER GSM2845325   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845325 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina MiSeq   SRP124609     AR07_S7.fastq.gz fastq 178903200.0 1064900.0 GSM2845325 r1                       SRX3374339 SRS2671567 SRA629220 GEO Broad Institute 1 1e-05   0.0   1.0       168   T   under 1.2% mapping rate illumina miseq unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44684 44684 SRR6268164 SRX3374332 SRS2671561 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   biorep A+ 10h.2 GSM2845318   source name:zebrafish embryos|developmental stage:10hpf|tissue:embryo biorep A+ 10h.2 Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:10hpf|tissue:embryo GSM2845318 GSM2845318: biorep A+ 10h.2; Danio rerio; OTHER GSM2845318   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845318 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina MiSeq   SRP124609     10hpf_S12.fastq.gz fastq 226568960.0 1416056.0 GSM2845318 r1 0:160 1:0 A:74664254;C:48098265;G:40952551;T:62847458;N:6432 160 0     74664254 48098265 40952551 62847458 6432 SRX3374332 SRS2671561 SRA629220 GEO Broad Institute 1 0.00023   2e-05   0.99987   0.87179   160   T   under 1.2% mapping rate illumina miseq unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44685 44685 SRR6268163 SRX3374331 SRS2671559 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   biorep A+ 8h.2 GSM2845317   source name:zebrafish embryos|developmental stage:8hpf|tissue:embryo biorep A+ 8h.2 Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:8hpf|tissue:embryo GSM2845317 GSM2845317: biorep A+ 8h.2; Danio rerio; OTHER GSM2845317   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845317 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina MiSeq   SRP124609     8hpf_S11.fastq.gz fastq 277302720.0 1733142.0 GSM2845317 r1 0:160 1:0 A:90214960;C:59749320;G:50838102;T:76483858;N:16480 160 0     90214960 59749320 50838102 76483858 16480 SRX3374331 SRS2671559 SRA629220 GEO Broad Institute 1 3e-05   0.0   0.99995   0.8   160   T   under 1.2% mapping rate illumina miseq unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44686 44686 SRR6268162 SRX3374330 SRS2671558 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   biorep A+ 6h.2 GSM2845316   source name:zebrafish embryos|developmental stage:6hpf|tissue:embryo biorep A+ 6h.2 Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:6hpf|tissue:embryo GSM2845316 GSM2845316: biorep A+ 6h.2; Danio rerio; OTHER GSM2845316   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845316 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina MiSeq   SRP124609     6hpf_S10.fastq.gz fastq 347336640.0 2170854.0 GSM2845316 r1 0:160 1:0 A:112549994;C:74770453;G:63347823;T:96645301;N:23069 160 0     112549994 74770453 63347823 96645301 23069 SRX3374330 SRS2671558 SRA629220 GEO Broad Institute 1 2e-05   0.0   0.99997   1.0   160   T   under 1.2% mapping rate illumina miseq unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44689 44689 SRR6268159 SRX3374327 SRS2671555 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   biorep A+ 10h.1 GSM2845313   source name:zebrafish embryos|developmental stage:10hpf|tissue:embryo biorep A+ 10h.1 Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:10hpf|tissue:embryo GSM2845313 GSM2845313: biorep A+ 10h.1; Danio rerio; OTHER GSM2845313   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845313 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina MiSeq   SRP124609     10hpf_S5.fastq.gz fastq 227268480.0 1420428.0 GSM2845313 r1 0:160 1:0 A:75021512;C:48442427;G:40892901;T:62896619;N:15021 160 0     75021512 48442427 40892901 62896619 15021 SRX3374327 SRS2671555 SRA629220 GEO Broad Institute 1 9e-05   0.0   0.99991   0.13333   160   T   under 1.2% mapping rate illumina miseq unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44690 44690 SRR6268158 SRX3374326 SRS2671554 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   biorep A+ 8h.1 GSM2845312   source name:zebrafish embryos|developmental stage:8hpf|tissue:embryo biorep A+ 8h.1 Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:8hpf|tissue:embryo GSM2845312 GSM2845312: biorep A+ 8h.1; Danio rerio; OTHER GSM2845312   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845312 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina MiSeq   SRP124609     8hpf_S4.fastq.gz fastq 291498560.0 1821866.0 GSM2845312 r1 0:160 1:0 A:95405630;C:62095343;G:53049839;T:80929783;N:17965 160 0     95405630 62095343 53049839 80929783 17965 SRX3374326 SRS2671554 SRA629220 GEO Broad Institute 1 1e-05   0.0   0.99997   1.0   160   T   under 1.2% mapping rate illumina miseq unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44691 44691 SRR6268157 SRX3374325 SRS2671553 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   biorep A+ 6h.1 GSM2845311   source name:zebrafish embryos|developmental stage:6hpf|tissue:embryo biorep A+ 6h.1 Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:6hpf|tissue:embryo GSM2845311 GSM2845311: biorep A+ 6h.1; Danio rerio; OTHER GSM2845311   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845311 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina MiSeq   SRP124609     6hpf_S3.fastq.gz fastq 344362400.0 2152265.0 GSM2845311 r1 0:160 1:0 A:111456568;C:74861160;G:63336980;T:94686986;N:20706 160 0     111456568 74861160 63336980 94686986 20706 SRX3374325 SRS2671553 SRA629220 GEO Broad Institute 1 3e-05   0.0   0.99997   1.0   160   T   under 1.2% mapping rate illumina miseq unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44695 44695 SRR6268153 SRX3374321 SRS2671548 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   embryo A 10h GSM2845307   source name:zebrafish embryos|developmental stage:10hpf|tissue:embryo embryo A 10h Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:10hpf|tissue:embryo GSM2845307 GSM2845307: embryo A 10h; Danio rerio; OTHER GSM2845307   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845307 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2500   SRP124609     SPE_0xA_AR10.fastq.gz fastq 198697400.0 1986974.0 GSM2845307 r1 0:100 A:67804840;C:37625181;G:37716734;T:55503500;N:47145 100       67804840 37625181 37716734 55503500 47145 SRX3374321 SRS2671548 SRA629220 GEO Broad Institute 1 4e-05   1e-05   0.99991   0.5   100   T   under 1.2% mapping rate illumina hiseq_era unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44696 44696 SRR6268152 SRX3374320 SRS2671550 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   embryo A 8h GSM2845306   source name:zebrafish embryos|developmental stage:8hpf|tissue:embryo embryo A 8h Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:8hpf|tissue:embryo GSM2845306 GSM2845306: embryo A 8h; Danio rerio; OTHER GSM2845306   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845306 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2500   SRP124609     SPE_0xA_AR09.fastq.gz fastq 392836100.0 3928361.0 GSM2845306 r1 0:100 A:137386195;C:71809820;G:72324431;T:111220224;N:95430 100       137386195 71809820 72324431 111220224 95430 SRX3374320 SRS2671550 SRA629220 GEO Broad Institute 1 3e-05   0.0   0.99991   0.0   100   T   under 1.2% mapping rate illumina hiseq_era unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44697 44697 SRR6268151 SRX3374319 SRS2671549 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   embryo A 7h GSM2845305   source name:zebrafish embryos|developmental stage:7hpf|tissue:embryo embryo A 7h Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:7hpf|tissue:embryo GSM2845305 GSM2845305: embryo A 7h; Danio rerio; OTHER GSM2845305   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845305 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2500   SRP124609     SPE_0xA_AR08.fastq.gz fastq 317338500.0 3173385.0 GSM2845305 r1 0:100 A:111088522;C:57862225;G:58342901;T:89968791;N:76061 100       111088522 57862225 58342901 89968791 76061 SRX3374319 SRS2671549 SRA629220 GEO Broad Institute 1 3e-05   0.0   0.99993   0.25   100   T   under 1.2% mapping rate illumina hiseq_era unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44698 44698 SRR6268150 SRX3374318 SRS2671547 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   embryo A 6h GSM2845304   source name:zebrafish embryos|developmental stage:6hpf|tissue:embryo embryo A 6h Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:6hpf|tissue:embryo GSM2845304 GSM2845304: embryo A 6h; Danio rerio; OTHER GSM2845304   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845304 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2500   SRP124609     SPE_0xA_AR07.fastq.gz fastq 261732400.0 2617324.0 GSM2845304 r1 0:100 A:87320893;C:49675297;G:50500129;T:74173127;N:62954 100       87320893 49675297 50500129 74173127 62954 SRX3374318 SRS2671547 SRA629220 GEO Broad Institute 1 2e-05   0.0   0.99995   0.0   100   T   under 1.2% mapping rate illumina hiseq_era unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44705 44705 SRR6268143 SRX3374311 SRS2671597 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   embryo A+ 10h GSM2845297   source name:zebrafish embryos|developmental stage:10hpf|tissue:embryo embryo A+ 10h Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:10hpf|tissue:embryo GSM2845297 GSM2845297: embryo A+ 10h; Danio rerio; OTHER GSM2845297   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845297 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2500   SRP124609     SPE_40xA_AR10.fastq.gz fastq 275983100.0 2759831.0 GSM2845297 r1 0:100 A:91324521;C:53348020;G:54834469;T:76419491;N:56599 100       91324521 53348020 54834469 76419491 56599 SRX3374311 SRS2671597 SRA629220 GEO Broad Institute 1 3e-05   0.0   0.99993   0.66666   100   T   under 1.2% mapping rate illumina hiseq_era unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44706 44706 SRR6268142 SRX3374310 SRS2671540 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   embryo A+ 8h GSM2845296   source name:zebrafish embryos|developmental stage:8hpf|tissue:embryo embryo A+ 8h Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:8hpf|tissue:embryo GSM2845296 GSM2845296: embryo A+ 8h; Danio rerio; OTHER GSM2845296   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845296 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2500   SRP124609     SPE_40xA_AR09.fastq.gz fastq 356804800.0 3568048.0 GSM2845296 r1 0:100 A:119536404;C:67554394;G:70133466;T:99507067;N:73469 100       119536404 67554394 70133466 99507067 73469 SRX3374310 SRS2671540 SRA629220 GEO Broad Institute 1 5e-05   0.0   0.99983   0.375   100   T   under 1.2% mapping rate illumina hiseq_era unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44707 44707 SRR6268141 SRX3374309 SRS2671538 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   embryo A+ 7h GSM2845295   source name:zebrafish embryos|developmental stage:7hpf|tissue:embryo embryo A+ 7h Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:7hpf|tissue:embryo GSM2845295 GSM2845295: embryo A+ 7h; Danio rerio; OTHER GSM2845295   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845295 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2500   SRP124609     SPE_40xA_AR08.fastq.gz fastq 443412400.0 4434124.0 GSM2845295 r1 0:100 A:148620758;C:84248870;G:86729304;T:123722045;N:91423 100       148620758 84248870 86729304 123722045 91423 SRX3374309 SRS2671538 SRA629220 GEO Broad Institute 1 9e-05   1e-05   0.99975   0.35714   100   T   under 1.2% mapping rate illumina hiseq_era unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
44708 44708 SRR6268140 SRX3374308 SRS2671537 SRP124609 PRJNA417597 Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation GSE106677 Other The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were coll…   pubmed:29225039   embryo A+ 6h GSM2845294   source name:zebrafish embryos|developmental stage:6hpf|tissue:embryo embryo A+ 6h Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5’ terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels zebrafish embryos Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage. Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3’UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3’UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3’UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage. developmental stage:6hpf|tissue:embryo GSM2845294 GSM2845294: embryo A+ 6h; Danio rerio; OTHER GSM2845294   1 Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors GEO Accession:GSM2845294 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2500   SRP124609     SPE_40xA_AR07.fastq.gz fastq 503622700.0 5036227.0 GSM2845294 r1 0:100 A:166949314;C:96806940;G:99443944;T:140317618;N:104884 100       166949314 96806940 99443944 140317618 104884 SRX3374308 SRS2671537 SRA629220 GEO Broad Institute 1 4e-05   0.0   0.99987   0.66666   100   T   under 1.2% mapping rate illumina hiseq_era unknown random_priming unknown bulk unknown unknown   United States 2017-11-08 Gastrula Embryo Embryo Imprecise All anatomical structures
49555 49555 SRR7942630 SRX4776895 SRS3857435 SRP162875 PRJNA493829 m5C profiles in zebrafish embryo sample. GSE120645 Other Total RNA was isolated from zebrafish embryos harvested at different stages using TRIzol® Reagent Ambion. mRNA was extracted with using Dynabeads® mRNA Purification Kit Ambion and subjected to TURBO™ DNase Invitrogen treatment at 37°C for 30 min and ethanol precipitation. Thus purified mRNA was used for RNA BisSeq library construction. Around 200 ng mRNA premixed with the in vitro transcribed Dhfr mRNA at a ratio of 300:1 Dhfr mRNA serves as methylation conversion control was fragmented to 100 nt fragments for 1 min at 90°C in 10× RNA Fragmentation Reagent Ambion then stopped by 10× RNA stop solution Ambion and precipitated with 100% ethanol. The RNA pellet was resuspended in 100 µl bisulfite solution pH 5.1 which is a 100:1 mixture of 40% sodium bisulfite Sigma and 600 µM hydroquinone Sigma and subjected to heat incubation at 75°C for 4.5 h. The reaction mixture was desalted by passing through Nanosep with 3K Omega 500/pk columns PALL Corporation with centrifugation. post washed with nuclease free water and centrifuged for five times the RNA was finally disolved in 75 µl nuclease free water and then desulfonated by incubation with an equal volume of 1 M Tris HCl pH 9.0 at 75 °C for 1 h. post ethanol precipitation the RNA was resuspended in 11 µl of RNase free water and subjected to library construction. Reverse transcription was carried out with superscript II Reverse Transcriptase Invitrogen and ACT random hexamers. The following procedures were performed with the KAPA Stranded mRNA Seq Kit KAPA according to the manufacturer's instructions.Libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 125 bp. Overall design: Examination of m5C levels in zebrafish embryo.       WT 6hpf m5C rep2 GSM3406896   source name:Zebrafish embryo|strain:AB strain|cell type:Zebrafish embryo|age:6 hpf|tissue:whole embryo WT 6hpf m5C rep2 Reads were aligned to the zv9 and hg19 genome assembly using meRanTK v1.2.0 m5C sites were called by meRanCall v1.2.0 and annotated by applying BEDTools’ intersectBed. Genome build: zv9 and hg19 Supplementary files format and content: m5C sites in two biological replicates. Zebrafish embryo   Total RNA was isolated from zebrafish embryos harvested at different stages using TRIzol® Reagent Ambion. mRNA was extracted with using Dynabeads® mRNA Purification Kit Ambion and subjected to TURBO™ DNase Invitrogen treatment at 37°C for 30 min and ethanol precipitation. Thus purified mRNA was used for RNA BisSeq library construction. Thus purified mRNA was used for RNA BisSeq library construction. Around 200 ng mRNA premixed with the in vitro transcribed Dhfr mRNA at a ratio of 300:1 Dhfr mRNA serves as methylation conversion control was fragmented to 100 nt fragments for 1 min at 90°C in 10× RNA Fragmentation Reagent Ambion then stopped by 10× RNA stop solution Ambion and precipitated with 100% ethanol. The RNA pellet was resuspended in 100 µl bisulfite solution pH 5.1 which is a 100:1 mixture of 40% sodium bisulfite Sigma and 600 µM hydroquinone Sigma and subjected to heat incubation at 75°C for 4.5 h. The reaction mixture was desalted by passing through Nanosep with 3K Omega 500/pk columns PALL Corporation with centrifugation. post washed with nuclease free water and centrifuged for five times the RNA was finally disolved in 75 μl nuclease free water and then desulfonated by incubation with an equal volume of 1 M Tris HCl pH 9.0 at 75 °C for 1 h. post ethanol precipitation the RNA was resuspended in 11 µl of RNase free water and subjected to library construction. Reverse transcription was carried out with superscript II Reverse Transcriptase Invitrogen and ACT random hexamers. The following procedures were performed with the KAPA Stranded mRNA Seq Kit KAPA according to the manufacturer’s instructions.Libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 125 bp. RNA BisSeq   strain:AB strain|cell type:Zebrafish embryo|age:6 hpf|tissue:whole embryo GSM3406896 GSM3406896: WT 6hpf m5C rep2; Danio rerio; OTHER GSM3406896   1 Total RNA was isolated from zebrafish embryos harvested at different stages using TRIzol® Reagent Ambion. mRNA was extracted with using Dynabeads® mRNA Purification Kit Ambion and subjected to TURBO™ DNase Invitrogen treatment at 37°C for 30 min and ethanol precipitation. Thus purified mRNA was used for RNA BisSeq library construction. Thus purified mRNA was used for RNA BisSeq library construction. Around 200 ng mRNA premixed with the in vitro transcribed Dhfr mRNA at a ratio of 300:1 Dhfr mRNA serves as methylation conversion control was fragmented to 100 nt fragments for 1 min at 90°C in 10× RNA Fragmentation Reagent Ambion then stopped by 10× RNA stop solution Ambion and precipitated with 100% ethanol. The RNA pellet was resuspended in 100 µl bisulfite solution pH 5.1 which is a 100:1 mixture of 40% sodium bisulfite Sigma and 600 µM hydroquinone Sigma and subjected to heat incubation at 75°C for 4.5 h. The reaction mixture was desalted by passing through Nanosep with 3K Omega 500/pk columns PALL Corporation with centrifugation. post washed with nuclease free water and centrifuged for five times the RNA was finally disolved in 75 μl nuclease free water and then desulfonated by incubation with an equal volume of 1 M Tris HCl pH 9.0 at 75 °C for 1 h. post ethanol precipitation the RNA was resuspended in 11 µl of RNase free water and subjected to library construction. Reverse transcription was carried out with superscript II Reverse Transcriptase Invitrogen and ACT random hexamers. The following procedures were performed with the KAPA Stranded mRNA Seq Kit KAPA according to the manufacturer's instructions.Libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 125 bp. RNA BisSeq GEO Accession:GSM3406896 OTHER TRANSCRIPTOMIC other PAIRED ILLUMINA Illumina HiSeq 2500   SRP162875     WT_6hpf_rep2_m5C_R1.fastq.gz WT_6hpf_rep2_m5C_R2.fastq.gz fastq fastq 19835440000.0 79341760.0 GSM3406896 r1 0:125 1:125 A:6759870592;C:3198281670;G:3585072759;T:6287844418;N:4370561 125 125     6759870592 3198281670 3585072759 6287844418 4370561 SRX4776895 SRS3857435 SRA786937 GEO Beijing Institute of Genomics (BIG) of Chinese Academy of Sciences (CAS) 2 0.00127 0.00115 0.00035 0.00032 0.9987 0.99882 0.73913 0.76712 125 125 T T mates < 9% mapping rate illumina hiseq_era unknown random_priming unknown bulk unknown unknown   China 2018-09-28 Gastrula Embryo Whole Organism All anatomical structures
49556 49556 SRR7942629 SRX4776894 SRS3857434 SRP162875 PRJNA493829 m5C profiles in zebrafish embryo sample. GSE120645 Other Total RNA was isolated from zebrafish embryos harvested at different stages using TRIzol® Reagent Ambion. mRNA was extracted with using Dynabeads® mRNA Purification Kit Ambion and subjected to TURBO™ DNase Invitrogen treatment at 37°C for 30 min and ethanol precipitation. Thus purified mRNA was used for RNA BisSeq library construction. Around 200 ng mRNA premixed with the in vitro transcribed Dhfr mRNA at a ratio of 300:1 Dhfr mRNA serves as methylation conversion control was fragmented to 100 nt fragments for 1 min at 90°C in 10× RNA Fragmentation Reagent Ambion then stopped by 10× RNA stop solution Ambion and precipitated with 100% ethanol. The RNA pellet was resuspended in 100 µl bisulfite solution pH 5.1 which is a 100:1 mixture of 40% sodium bisulfite Sigma and 600 µM hydroquinone Sigma and subjected to heat incubation at 75°C for 4.5 h. The reaction mixture was desalted by passing through Nanosep with 3K Omega 500/pk columns PALL Corporation with centrifugation. post washed with nuclease free water and centrifuged for five times the RNA was finally disolved in 75 µl nuclease free water and then desulfonated by incubation with an equal volume of 1 M Tris HCl pH 9.0 at 75 °C for 1 h. post ethanol precipitation the RNA was resuspended in 11 µl of RNase free water and subjected to library construction. Reverse transcription was carried out with superscript II Reverse Transcriptase Invitrogen and ACT random hexamers. The following procedures were performed with the KAPA Stranded mRNA Seq Kit KAPA according to the manufacturer's instructions.Libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 125 bp. Overall design: Examination of m5C levels in zebrafish embryo.       WT 6hpf m5C rep1 GSM3406895   source name:Zebrafish embryo|strain:AB strain|cell type:Zebrafish embryo|age:6 hpf|tissue:whole embryo WT 6hpf m5C rep1 Reads were aligned to the zv9 and hg19 genome assembly using meRanTK v1.2.0 m5C sites were called by meRanCall v1.2.0 and annotated by applying BEDTools’ intersectBed. Genome build: zv9 and hg19 Supplementary files format and content: m5C sites in two biological replicates. Zebrafish embryo   Total RNA was isolated from zebrafish embryos harvested at different stages using TRIzol® Reagent Ambion. mRNA was extracted with using Dynabeads® mRNA Purification Kit Ambion and subjected to TURBO™ DNase Invitrogen treatment at 37°C for 30 min and ethanol precipitation. Thus purified mRNA was used for RNA BisSeq library construction. Thus purified mRNA was used for RNA BisSeq library construction. Around 200 ng mRNA premixed with the in vitro transcribed Dhfr mRNA at a ratio of 300:1 Dhfr mRNA serves as methylation conversion control was fragmented to 100 nt fragments for 1 min at 90°C in 10× RNA Fragmentation Reagent Ambion then stopped by 10× RNA stop solution Ambion and precipitated with 100% ethanol. The RNA pellet was resuspended in 100 µl bisulfite solution pH 5.1 which is a 100:1 mixture of 40% sodium bisulfite Sigma and 600 µM hydroquinone Sigma and subjected to heat incubation at 75°C for 4.5 h. The reaction mixture was desalted by passing through Nanosep with 3K Omega 500/pk columns PALL Corporation with centrifugation. post washed with nuclease free water and centrifuged for five times the RNA was finally disolved in 75 μl nuclease free water and then desulfonated by incubation with an equal volume of 1 M Tris HCl pH 9.0 at 75 °C for 1 h. post ethanol precipitation the RNA was resuspended in 11 µl of RNase free water and subjected to library construction. Reverse transcription was carried out with superscript II Reverse Transcriptase Invitrogen and ACT random hexamers. The following procedures were performed with the KAPA Stranded mRNA Seq Kit KAPA according to the manufacturer’s instructions.Libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 125 bp. RNA BisSeq   strain:AB strain|cell type:Zebrafish embryo|age:6 hpf|tissue:whole embryo GSM3406895 GSM3406895: WT 6hpf m5C rep1; Danio rerio; OTHER GSM3406895   1 Total RNA was isolated from zebrafish embryos harvested at different stages using TRIzol® Reagent Ambion. mRNA was extracted with using Dynabeads® mRNA Purification Kit Ambion and subjected to TURBO™ DNase Invitrogen treatment at 37°C for 30 min and ethanol precipitation. Thus purified mRNA was used for RNA BisSeq library construction. Thus purified mRNA was used for RNA BisSeq library construction. Around 200 ng mRNA premixed with the in vitro transcribed Dhfr mRNA at a ratio of 300:1 Dhfr mRNA serves as methylation conversion control was fragmented to 100 nt fragments for 1 min at 90°C in 10× RNA Fragmentation Reagent Ambion then stopped by 10× RNA stop solution Ambion and precipitated with 100% ethanol. The RNA pellet was resuspended in 100 µl bisulfite solution pH 5.1 which is a 100:1 mixture of 40% sodium bisulfite Sigma and 600 µM hydroquinone Sigma and subjected to heat incubation at 75°C for 4.5 h. The reaction mixture was desalted by passing through Nanosep with 3K Omega 500/pk columns PALL Corporation with centrifugation. post washed with nuclease free water and centrifuged for five times the RNA was finally disolved in 75 μl nuclease free water and then desulfonated by incubation with an equal volume of 1 M Tris HCl pH 9.0 at 75 °C for 1 h. post ethanol precipitation the RNA was resuspended in 11 µl of RNase free water and subjected to library construction. Reverse transcription was carried out with superscript II Reverse Transcriptase Invitrogen and ACT random hexamers. The following procedures were performed with the KAPA Stranded mRNA Seq Kit KAPA according to the manufacturer's instructions.Libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 125 bp. RNA BisSeq GEO Accession:GSM3406895 OTHER TRANSCRIPTOMIC other PAIRED ILLUMINA Illumina HiSeq 2500   SRP162875     WT_6hpf_rep1_m5C_R1.fastq.gz WT_6hpf_rep1_m5C_R2.fastq.gz fastq fastq 22436339250.0 89745357.0 GSM3406895 r1 0:125 1:125 A:7538129383;C:3757045353;G:4234311296;T:6842033064;N:64820154 125 125     7538129383 3757045353 4234311296 6842033064 64820154 SRX4776894 SRS3857434 SRA786937 GEO Beijing Institute of Genomics (BIG) of Chinese Academy of Sciences (CAS) 2 0.00111 0.00098 0.00035 0.0003 0.99813 0.99837 0.47183 0.44531 125 125 T T mates < 9% mapping rate illumina hiseq_era unknown random_priming unknown bulk unknown unknown   China 2018-09-28 Gastrula Embryo Whole Organism All anatomical structures
49565 49565 SRR11537844 SRX8108899 SRS6473985 SRP163087 PRJNA494251 Dynamic landscapes and regulome of mRNA structure during zebrafish early embryogenesis GSE120724 Other We resolved the RNA secondary structure during zebrafish early embryogenesis based on in vivo click selective 2' hydroxyl acylation and profiling experiment icSHAPE. We analyzed the RNA structure dynamics among different development stages. Also we studied which factors regulate maternal gene decay by RNA structure switch. Overall design: icSHAPE of 12 samples from zebrafish embryos at six different early development stages and 4 samples from Elavl1a deficient zebrafish embryos at sphere and shield stages including two treated with DMSO as control and two treated with NAIN3 for each stage or condition. RNA seq of 6 samples from zebrafish embryos at 2 4 6 h.p.f. and 4 samples from Elavl1a deficient zebrafish embryos at 4 and 6 h.p.f. including two biological replicates. iCLIP seq of 4 samples from flag elavl1a mRNA injected zebrafish embryos at xxx and 6 h.p.f. including two biological replicates.   pubmed:32423473   Shield Elavl1a MO NAIN3.rep2 GSM4473292   tissue:Shield Elavl1a MO NAIN3|strain background:AB|age:6hpf|cell type:elavl1a morphants zebrafish embryos cells|treatment:NAI N3 Shield Elavl1a MO NAIN3.rep2 Library strategy: icSHAPE trimmed three prime adaptor from reads by cutadapt trimmed low quality bases by trimmomatic collapsed duplicated reads as icSHAPE pipeline trimmed the leading 13nt UMI mapping reads to zebrafish reference transcriptomez10 by bowtie2 using the same parameters in icSHAPE pipeline calculate rpkm as icshape pipeline calulate reverse transcription stop sitesRT stop as icshape pipeline RT stop was normalized by using sliding window then icSHAPE score was calculated as icshape pipeline Genome build: z10 Supplementary files format and content: tab delimited text files for icshape files. First column is the Refseq ID second is transcript length third is RPKM then every column is the nucleotide resolution icSHAPE score. NULL means no confident score. Tab delimited text files for read counts and rpkm files. First column is the Refseq ID second is transcript length third is the unique mapped reads count fourth is the mulitple mapped reads count fifth is the RPKM. Shield Elavl1a MO NAIN3 For Elavl1a morphants generation morpholino oligonucleotides of elavl1a were injected into one cell stage embryos then embryos were harvested at desired stages. For in vivo treatment cells was incubated with NAI N3 or DMSO at 28.5°C for 5min The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol® Reagent invitrogen.To enrich polyA RNA the total RNA was subjected to polyA selection. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37℃ for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/µl RiboLock 0.1 U/µl FastAP Life Technology 2 U/µl T4 PNKNEB before 3’ adapter ligation with addition of 10 µl ligation mix 5mM DTT 0.5µM 5’ adenylated and 3’ blocked linker 3’ bio for unmodified 3’ ddc for modified 0.66 U/µl T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25℃ for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1µl FastAP 0.2µl SSB Promega 0.8µl Ribolock and 1µl 5’ Deadenylase NEB in 1X NEB buffer 2 NEB at 30℃ for 90 min. And then 1µl RecJf NEB was added with another incubation at 37℃ for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq， Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer’s protocol. Zebrafish wild type strain AB was raised in system water at 28.5°C under standard conditions. strain background:AB|age:6hpf|cell type:elavl1a morphants zebrafish embryos cells|treatment:NAI N3 GSM4473292 GSM4473292: Shield Elavl1a MO NAIN3.rep2; Danio rerio; OTHER GSM4473292   1 The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol® Reagent invitrogen. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37℃ for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/µl RiboLock 0.1 U/µl FastAP Life Technology 2 U/µl T4 PNKNEB before three prime adapter ligation with addition of 10 µl ligation mix 5mM DTT 0.5µM five prime adenylated and three prime blocked linker three prime bio for unmodified three prime ddc for modified 0.66 U/µl T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25℃ for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1µl FastAP 0.2µl SSB Promega 0.8µl Ribolock and 1µl five prime Deadenylase NEB in 1X NEB buffer 2 NEB at 30℃ for 90 min. And then 1µl RecJf NEB was added with another incubation at 37℃ for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq， Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer's protocol. GEO Accession:GSM4473292 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2500   SRP163087     Shield_Elavl1a_MO_NAIN3.rep2.fastq.gz fastq 125703095100.0 838020634.0 GSM4473292 r1 0:150 1:0 A:32023181587;C:24765137819;G:46378114606;T:22535691781;N:969307 150 0     32023181587 24765137819 46378114606 22535691781 969307 SRX8108899 SRS6473985 SRA787572 GEO Life Science, Tsinghua University 1 0.11412   0.02142   0.92147   0.70154   150   B   usable mapping rate illumina hiseq_era unknown poly_a unknown bulk clip iclip   China 2020-04-12 Gastrula Embryo Embryo Imprecise All anatomical structures
49566 49566 SRR11537843 SRX8108898 SRS6473984 SRP163087 PRJNA494251 Dynamic landscapes and regulome of mRNA structure during zebrafish early embryogenesis GSE120724 Other We resolved the RNA secondary structure during zebrafish early embryogenesis based on in vivo click selective 2' hydroxyl acylation and profiling experiment icSHAPE. We analyzed the RNA structure dynamics among different development stages. Also we studied which factors regulate maternal gene decay by RNA structure switch. Overall design: icSHAPE of 12 samples from zebrafish embryos at six different early development stages and 4 samples from Elavl1a deficient zebrafish embryos at sphere and shield stages including two treated with DMSO as control and two treated with NAIN3 for each stage or condition. RNA seq of 6 samples from zebrafish embryos at 2 4 6 h.p.f. and 4 samples from Elavl1a deficient zebrafish embryos at 4 and 6 h.p.f. including two biological replicates. iCLIP seq of 4 samples from flag elavl1a mRNA injected zebrafish embryos at xxx and 6 h.p.f. including two biological replicates.   pubmed:32423473   Shield Elavl1a MO NAIN3.rep1 GSM4473291   tissue:Shield Elavl1a MO NAIN3|strain background:AB|age:6hpf|cell type:elavl1a morphants zebrafish embryos cells|treatment:NAI N3 Shield Elavl1a MO NAIN3.rep1 Library strategy: icSHAPE trimmed three prime adaptor from reads by cutadapt trimmed low quality bases by trimmomatic collapsed duplicated reads as icSHAPE pipeline trimmed the leading 13nt UMI mapping reads to zebrafish reference transcriptomez10 by bowtie2 using the same parameters in icSHAPE pipeline calculate rpkm as icshape pipeline calulate reverse transcription stop sitesRT stop as icshape pipeline RT stop was normalized by using sliding window then icSHAPE score was calculated as icshape pipeline Genome build: z10 Supplementary files format and content: tab delimited text files for icshape files. First column is the Refseq ID second is transcript length third is RPKM then every column is the nucleotide resolution icSHAPE score. NULL means no confident score. Tab delimited text files for read counts and rpkm files. First column is the Refseq ID second is transcript length third is the unique mapped reads count fourth is the mulitple mapped reads count fifth is the RPKM. Shield Elavl1a MO NAIN3 For Elavl1a morphants generation morpholino oligonucleotides of elavl1a were injected into one cell stage embryos then embryos were harvested at desired stages. For in vivo treatment cells was incubated with NAI N3 or DMSO at 28.5°C for 5min The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol® Reagent invitrogen.To enrich polyA RNA the total RNA was subjected to polyA selection. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37℃ for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/µl RiboLock 0.1 U/µl FastAP Life Technology 2 U/µl T4 PNKNEB before 3’ adapter ligation with addition of 10 µl ligation mix 5mM DTT 0.5µM 5’ adenylated and 3’ blocked linker 3’ bio for unmodified 3’ ddc for modified 0.66 U/µl T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25℃ for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1µl FastAP 0.2µl SSB Promega 0.8µl Ribolock and 1µl 5’ Deadenylase NEB in 1X NEB buffer 2 NEB at 30℃ for 90 min. And then 1µl RecJf NEB was added with another incubation at 37℃ for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq， Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer’s protocol. Zebrafish wild type strain AB was raised in system water at 28.5°C under standard conditions. strain background:AB|age:6hpf|cell type:elavl1a morphants zebrafish embryos cells|treatment:NAI N3 GSM4473291 GSM4473291: Shield Elavl1a MO NAIN3.rep1; Danio rerio; OTHER GSM4473291   1 The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol® Reagent invitrogen. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37℃ for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/µl RiboLock 0.1 U/µl FastAP Life Technology 2 U/µl T4 PNKNEB before three prime adapter ligation with addition of 10 µl ligation mix 5mM DTT 0.5µM five prime adenylated and three prime blocked linker three prime bio for unmodified three prime ddc for modified 0.66 U/µl T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25℃ for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1µl FastAP 0.2µl SSB Promega 0.8µl Ribolock and 1µl five prime Deadenylase NEB in 1X NEB buffer 2 NEB at 30℃ for 90 min. And then 1µl RecJf NEB was added with another incubation at 37℃ for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq， Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer's protocol. GEO Accession:GSM4473291 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2500   SRP163087     Shield_Elavl1a_MO_NAIN3.rep1.fastq.gz fastq 97684476300.0 651229842.0 GSM4473291 r1 0:150 1:0 A:23448978329;C:19171932414;G:36104881456;T:18957926032;N:758069 150 0     23448978329 19171932414 36104881456 18957926032 758069 SRX8108898 SRS6473984 SRA787572 GEO Life Science, Tsinghua University 1 0.07298   0.01306   0.93789   0.71752   150   B   usable mapping rate illumina hiseq_era unknown poly_a unknown bulk clip iclip   China 2020-04-12 Gastrula Embryo Embryo Imprecise All anatomical structures
49567 49567 SRR11537842 SRX8108897 SRS6473983 SRP163087 PRJNA494251 Dynamic landscapes and regulome of mRNA structure during zebrafish early embryogenesis GSE120724 Other We resolved the RNA secondary structure during zebrafish early embryogenesis based on in vivo click selective 2' hydroxyl acylation and profiling experiment icSHAPE. We analyzed the RNA structure dynamics among different development stages. Also we studied which factors regulate maternal gene decay by RNA structure switch. Overall design: icSHAPE of 12 samples from zebrafish embryos at six different early development stages and 4 samples from Elavl1a deficient zebrafish embryos at sphere and shield stages including two treated with DMSO as control and two treated with NAIN3 for each stage or condition. RNA seq of 6 samples from zebrafish embryos at 2 4 6 h.p.f. and 4 samples from Elavl1a deficient zebrafish embryos at 4 and 6 h.p.f. including two biological replicates. iCLIP seq of 4 samples from flag elavl1a mRNA injected zebrafish embryos at xxx and 6 h.p.f. including two biological replicates.   pubmed:32423473   Shield Elavl1a MO DMSO.rep2 GSM4473290   tissue:Shield Elavl1a MO DMSO|strain background:AB|age:6hpf|cell type:elavl1a morphants zebrafish embryos cells|treatment:DMSO Shield Elavl1a MO DMSO.rep2 Library strategy: icSHAPE trimmed three prime adaptor from reads by cutadapt trimmed low quality bases by trimmomatic collapsed duplicated reads as icSHAPE pipeline trimmed the leading 13nt UMI mapping reads to zebrafish reference transcriptomez10 by bowtie2 using the same parameters in icSHAPE pipeline calculate rpkm as icshape pipeline calulate reverse transcription stop sitesRT stop as icshape pipeline RT stop was normalized by using sliding window then icSHAPE score was calculated as icshape pipeline Genome build: z10 Supplementary files format and content: tab delimited text files for icshape files. First column is the Refseq ID second is transcript length third is RPKM then every column is the nucleotide resolution icSHAPE score. NULL means no confident score. Tab delimited text files for read counts and rpkm files. First column is the Refseq ID second is transcript length third is the unique mapped reads count fourth is the mulitple mapped reads count fifth is the RPKM. Shield Elavl1a MO DMSO For Elavl1a morphants generation morpholino oligonucleotides of elavl1a were injected into one cell stage embryos then embryos were harvested at desired stages. For in vivo treatment cells was incubated with NAI N3 or DMSO at 28.5°C for 5min The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol® Reagent invitrogen.To enrich polyA RNA the total RNA was subjected to polyA selection. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37℃ for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/µl RiboLock 0.1 U/µl FastAP Life Technology 2 U/µl T4 PNKNEB before 3’ adapter ligation with addition of 10 µl ligation mix 5mM DTT 0.5µM 5’ adenylated and 3’ blocked linker 3’ bio for unmodified 3’ ddc for modified 0.66 U/µl T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25℃ for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1µl FastAP 0.2µl SSB Promega 0.8µl Ribolock and 1µl 5’ Deadenylase NEB in 1X NEB buffer 2 NEB at 30℃ for 90 min. And then 1µl RecJf NEB was added with another incubation at 37℃ for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq， Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer’s protocol. Zebrafish wild type strain AB was raised in system water at 28.5°C under standard conditions. strain background:AB|age:6hpf|cell type:elavl1a morphants zebrafish embryos cells|treatment:DMSO GSM4473290 GSM4473290: Shield Elavl1a MO DMSO.rep2; Danio rerio; OTHER GSM4473290   1 The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol® Reagent invitrogen. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37℃ for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/µl RiboLock 0.1 U/µl FastAP Life Technology 2 U/µl T4 PNKNEB before three prime adapter ligation with addition of 10 µl ligation mix 5mM DTT 0.5µM five prime adenylated and three prime blocked linker three prime bio for unmodified three prime ddc for modified 0.66 U/µl T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25℃ for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1µl FastAP 0.2µl SSB Promega 0.8µl Ribolock and 1µl five prime Deadenylase NEB in 1X NEB buffer 2 NEB at 30℃ for 90 min. And then 1µl RecJf NEB was added with another incubation at 37℃ for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq， Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer's protocol. GEO Accession:GSM4473290 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2500   SRP163087     Shield_Elavl1a_MO_DMSO.rep2.fastq.gz fastq 77964750450.0 519765003.0 GSM4473290 r1 0:150 1:0 A:23155551814;C:19934925592;G:15766394313;T:19078082759;N:29795972 150 0     23155551814 19934925592 15766394313 19078082759 29795972 SRX8108897 SRS6473983 SRA787572 GEO Life Science, Tsinghua University 1 0.07142   0.01724   0.94223   0.779   150   B   usable mapping rate illumina hiseq_era unknown poly_a unknown bulk clip iclip   China 2020-04-12 Gastrula Embryo Embryo Imprecise All anatomical structures
49568 49568 SRR11537841 SRX8108896 SRS6473982 SRP163087 PRJNA494251 Dynamic landscapes and regulome of mRNA structure during zebrafish early embryogenesis GSE120724 Other We resolved the RNA secondary structure during zebrafish early embryogenesis based on in vivo click selective 2' hydroxyl acylation and profiling experiment icSHAPE. We analyzed the RNA structure dynamics among different development stages. Also we studied which factors regulate maternal gene decay by RNA structure switch. Overall design: icSHAPE of 12 samples from zebrafish embryos at six different early development stages and 4 samples from Elavl1a deficient zebrafish embryos at sphere and shield stages including two treated with DMSO as control and two treated with NAIN3 for each stage or condition. RNA seq of 6 samples from zebrafish embryos at 2 4 6 h.p.f. and 4 samples from Elavl1a deficient zebrafish embryos at 4 and 6 h.p.f. including two biological replicates. iCLIP seq of 4 samples from flag elavl1a mRNA injected zebrafish embryos at xxx and 6 h.p.f. including two biological replicates.   pubmed:32423473   Shield Elavl1a MO DMSO.rep1 GSM4473289   tissue:Shield Elavl1a MO DMSO|strain background:AB|age:6hpf|cell type:elavl1a morphants zebrafish embryos cells|treatment:DMSO Shield Elavl1a MO DMSO.rep1 Library strategy: icSHAPE trimmed three prime adaptor from reads by cutadapt trimmed low quality bases by trimmomatic collapsed duplicated reads as icSHAPE pipeline trimmed the leading 13nt UMI mapping reads to zebrafish reference transcriptomez10 by bowtie2 using the same parameters in icSHAPE pipeline calculate rpkm as icshape pipeline calulate reverse transcription stop sitesRT stop as icshape pipeline RT stop was normalized by using sliding window then icSHAPE score was calculated as icshape pipeline Genome build: z10 Supplementary files format and content: tab delimited text files for icshape files. First column is the Refseq ID second is transcript length third is RPKM then every column is the nucleotide resolution icSHAPE score. NULL means no confident score. Tab delimited text files for read counts and rpkm files. First column is the Refseq ID second is transcript length third is the unique mapped reads count fourth is the mulitple mapped reads count fifth is the RPKM. Shield Elavl1a MO DMSO For Elavl1a morphants generation morpholino oligonucleotides of elavl1a were injected into one cell stage embryos then embryos were harvested at desired stages. For in vivo treatment cells was incubated with NAI N3 or DMSO at 28.5°C for 5min The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol® Reagent invitrogen.To enrich polyA RNA the total RNA was subjected to polyA selection. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37℃ for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/µl RiboLock 0.1 U/µl FastAP Life Technology 2 U/µl T4 PNKNEB before 3’ adapter ligation with addition of 10 µl ligation mix 5mM DTT 0.5µM 5’ adenylated and 3’ blocked linker 3’ bio for unmodified 3’ ddc for modified 0.66 U/µl T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25℃ for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1µl FastAP 0.2µl SSB Promega 0.8µl Ribolock and 1µl 5’ Deadenylase NEB in 1X NEB buffer 2 NEB at 30℃ for 90 min. And then 1µl RecJf NEB was added with another incubation at 37℃ for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq， Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer’s protocol. Zebrafish wild type strain AB was raised in system water at 28.5°C under standard conditions. strain background:AB|age:6hpf|cell type:elavl1a morphants zebrafish embryos cells|treatment:DMSO GSM4473289 GSM4473289: Shield Elavl1a MO DMSO.rep1; Danio rerio; OTHER GSM4473289   1 The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol® Reagent invitrogen. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37℃ for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/µl RiboLock 0.1 U/µl FastAP Life Technology 2 U/µl T4 PNKNEB before three prime adapter ligation with addition of 10 µl ligation mix 5mM DTT 0.5µM five prime adenylated and three prime blocked linker three prime bio for unmodified three prime ddc for modified 0.66 U/µl T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25℃ for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1µl FastAP 0.2µl SSB Promega 0.8µl Ribolock and 1µl five prime Deadenylase NEB in 1X NEB buffer 2 NEB at 30℃ for 90 min. And then 1µl RecJf NEB was added with another incubation at 37℃ for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq， Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer's protocol. GEO Accession:GSM4473289 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2500   SRP163087     Shield_Elavl1a_MO_DMSO.rep1.fastq.gz fastq 24430935750.0 162872905.0 GSM4473289 r1 0:150 1:0 A:6922313808;C:5215135549;G:7472148111;T:4821148458;N:189824 150 0     6922313808 5215135549 7472148111 4821148458 189824 SRX8108896 SRS6473982 SRA787572 GEO Life Science, Tsinghua University 1 0.27515   0.04834   0.86147   0.4542   150   B   usable mapping rate illumina hiseq_era unknown poly_a unknown bulk clip iclip   China 2020-04-12 Gastrula Embryo Embryo Imprecise All anatomical structures
49573 49573 SRR10434662 SRX7130632 SRS5639976 SRP163087 PRJNA494251 Dynamic landscapes and regulome of mRNA structure during zebrafish early embryogenesis GSE120724 Other We resolved the RNA secondary structure during zebrafish early embryogenesis based on in vivo click selective 2' hydroxyl acylation and profiling experiment icSHAPE. We analyzed the RNA structure dynamics among different development stages. Also we studied which factors regulate maternal gene decay by RNA structure switch. Overall design: icSHAPE of 12 samples from zebrafish embryos at six different early development stages and 4 samples from Elavl1a deficient zebrafish embryos at sphere and shield stages including two treated with DMSO as control and two treated with NAIN3 for each stage or condition. RNA seq of 6 samples from zebrafish embryos at 2 4 6 h.p.f. and 4 samples from Elavl1a deficient zebrafish embryos at 4 and 6 h.p.f. including two biological replicates. iCLIP seq of 4 samples from flag elavl1a mRNA injected zebrafish embryos at xxx and 6 h.p.f. including two biological replicates.   pubmed:32423473   shield Flag Elavl1a iCLIP rep2 GSM4157939   tissue:zebrafish embryos|strain:zebrafish embryos|cell type:normal zebrafish embryos cells|age:6hpf shield Flag Elavl1a iCLIP rep2 The proceesing procedure is similar to CTK toolhttps://zhanglab.c2b2.columbia.edu/index.php/ICLIP data analysis using CTK. First we trimmed three prime adaptor from reads by cutadapt collapsed duplicated reads by fastq2collapse.pl in CTK tool stripped 5’ degenerated barcode mapped clean reads to zebrafish reference genomez10 by bwav0.7.17. data from replicates was merged for peak calling We use CTK tool CITS mode to call the peaks. Reads were clustered using tag2cluster.pl with parameters big s maxgap " 1" peaks are identified using tag2peak.pl with parameters big ss v prefix "CITS" gap 25 p 0.05 Genome build: z10 Supplementary files format and content: iCLIP peaks identified by CTK tool were provided in bed file format. The first three columns represent the 0 based location of the peak in the genome and the last column represents the strand.The fourth column reprents the peak ID and describes the gene locus peak heightPH expected PH/backgroundPH0 and pvalueP. The fifth column represents the peak height. zebrafish embryos   iCLIP seq was carried out as previously described Despic et al. 2017. Breifly flag elavl1a mRNA injected embryos were collected at xxx h.p.f. and 6 h.p.f.. 400 zebrafish embryos were irradiated twice with 0.8 J/cm2 Stratalinker 2400 Stratagene lysed and subjected to mild RNA fragmentation. Crosslinked RNA proteins complexes were immunopurified using Anti FLAG M2 Magnetic Beads Merck M8823 for 4 h.p.f. and 6 h.p.f. at 4 °C. RNA was extracted Library construction was performed by using Smarter smRNA Seq kit Clontech Laboratories Inc Zebrafish wild type strain AB was raised in system water at 28.5°C under standard conditions. strain:zebrafish embryos|cell type:normal zebrafish embryos cells|age:6hpf GSM4157939 GSM4157939: shield Flag Elavl1a iCLIP rep2; Danio rerio; OTHER GSM4157939   1 iCLIP seq was carried out as previously described Despic et al. 2017. Breifly flag elavl1a mRNA injected embryos were collected at xxx h.p.f. and 6 h.p.f.. 400 zebrafish embryos were irradiated twice with 0.8 J/cm2 Stratalinker 2400 Stratagene lysed and subjected to mild RNA fragmentation. Crosslinked RNA proteins complexes were immunopurified using Anti FLAG M2 Magnetic Beads Merck M8823 for 4 h.p.f. and 6 h.p.f. at 4 °C. RNA was extracted Library construction was performed by using Smarter smRNA Seq kit Clontech Laboratories Inc GEO Accession:GSM4157939 OTHER TRANSCRIPTOMIC other PAIRED ILLUMINA HiSeq X Ten   SRP163087     Shield_Elavl1a_iCLIP_rep2.R2.fq.gz Shield_Elavl1a_iCLIP_rep2.R1.fq.gz fastq fastq 23879342400.0 79597808.0 GSM4157939 r1 0:150 1:150 A:6335254303;C:5166953187;G:6017120995;T:6358132998;N:1880917 150 150     6335254303 5166953187 6017120995 6358132998 1880917 SRX7130632 SRS5639976 SRA787572 GEO Life Science, Tsinghua University 2 0.14294 0.15151 0.03583 0.08959 0.99397 0.98269 0.96727 0.75507 150 150 B B biological fallback assumption illumina hiseq_era full_length poly_a smarter bulk clip iclip   China 2019-11-12 Gastrula Embryo Embryo Imprecise All anatomical structures
49574 49574 SRR10434661 SRX7130631 SRS5639975 SRP163087 PRJNA494251 Dynamic landscapes and regulome of mRNA structure during zebrafish early embryogenesis GSE120724 Other We resolved the RNA secondary structure during zebrafish early embryogenesis based on in vivo click selective 2' hydroxyl acylation and profiling experiment icSHAPE. We analyzed the RNA structure dynamics among different development stages. Also we studied which factors regulate maternal gene decay by RNA structure switch. Overall design: icSHAPE of 12 samples from zebrafish embryos at six different early development stages and 4 samples from Elavl1a deficient zebrafish embryos at sphere and shield stages including two treated with DMSO as control and two treated with NAIN3 for each stage or condition. RNA seq of 6 samples from zebrafish embryos at 2 4 6 h.p.f. and 4 samples from Elavl1a deficient zebrafish embryos at 4 and 6 h.p.f. including two biological replicates. iCLIP seq of 4 samples from flag elavl1a mRNA injected zebrafish embryos at xxx and 6 h.p.f. including two biological replicates.   pubmed:32423473   shield Flag Elavl1a iCLIP rep1 GSM4157938   tissue:zebrafish embryos|strain:zebrafish embryos|cell type:normal zebrafish embryos cells|age:6hpf shield Flag Elavl1a iCLIP rep1 The proceesing procedure is similar to CTK toolhttps://zhanglab.c2b2.columbia.edu/index.php/ICLIP data analysis using CTK. First we trimmed three prime adaptor from reads by cutadapt collapsed duplicated reads by fastq2collapse.pl in CTK tool stripped 5’ degenerated barcode mapped clean reads to zebrafish reference genomez10 by bwav0.7.17. data from replicates was merged for peak calling We use CTK tool CITS mode to call the peaks. Reads were clustered using tag2cluster.pl with parameters big s maxgap " 1" peaks are identified using tag2peak.pl with parameters big ss v prefix "CITS" gap 25 p 0.05 Genome build: z10 Supplementary files format and content: iCLIP peaks identified by CTK tool were provided in bed file format. The first three columns represent the 0 based location of the peak in the genome and the last column represents the strand.The fourth column reprents the peak ID and describes the gene locus peak heightPH expected PH/backgroundPH0 and pvalueP. The fifth column represents the peak height. zebrafish embryos   iCLIP seq was carried out as previously described Despic et al. 2017. Breifly flag elavl1a mRNA injected embryos were collected at xxx h.p.f. and 6 h.p.f.. 400 zebrafish embryos were irradiated twice with 0.8 J/cm2 Stratalinker 2400 Stratagene lysed and subjected to mild RNA fragmentation. Crosslinked RNA proteins complexes were immunopurified using Anti FLAG M2 Magnetic Beads Merck M8823 for 4 h.p.f. and 6 h.p.f. at 4 °C. RNA was extracted Library construction was performed by using Smarter smRNA Seq kit Clontech Laboratories Inc Zebrafish wild type strain AB was raised in system water at 28.5°C under standard conditions. strain:zebrafish embryos|cell type:normal zebrafish embryos cells|age:6hpf GSM4157938 GSM4157938: shield Flag Elavl1a iCLIP rep1; Danio rerio; OTHER GSM4157938   1 iCLIP seq was carried out as previously described Despic et al. 2017. Breifly flag elavl1a mRNA injected embryos were collected at xxx h.p.f. and 6 h.p.f.. 400 zebrafish embryos were irradiated twice with 0.8 J/cm2 Stratalinker 2400 Stratagene lysed and subjected to mild RNA fragmentation. Crosslinked RNA proteins complexes were immunopurified using Anti FLAG M2 Magnetic Beads Merck M8823 for 4 h.p.f. and 6 h.p.f. at 4 °C. RNA was extracted Library construction was performed by using Smarter smRNA Seq kit Clontech Laboratories Inc GEO Accession:GSM4157938 OTHER TRANSCRIPTOMIC other PAIRED ILLUMINA HiSeq X Ten   SRP163087     Shield_Elavl1a_iCLIP_rep1.R1.fq.gz Shield_Elavl1a_iCLIP_rep1.R2.fq.gz fastq fastq 22375977600.0 74586592.0 GSM4157938 r1 0:150 1:150 A:6243025236;C:5008499337;G:5418170547;T:5704514909;N:1767571 150 150     6243025236 5008499337 5418170547 5704514909 1767571 SRX7130631 SRS5639975 SRA787572 GEO Life Science, Tsinghua University 2 0.05144 0.12625 0.01219 0.09577 0.99734 0.98113 0.8629 0.52429 150 150 B B mate1-mate2 similar by mapping diff illumina hiseq_era full_length poly_a smarter bulk clip iclip   China 2019-11-12 Gastrula Embryo Embryo Imprecise All anatomical structures
49587 49587 SRR7947911 SRX4781863 SRS3862055 SRP163087 PRJNA494251 Dynamic landscapes and regulome of mRNA structure during zebrafish early embryogenesis GSE120724 Other We resolved the RNA secondary structure during zebrafish early embryogenesis based on in vivo click selective 2' hydroxyl acylation and profiling experiment icSHAPE. We analyzed the RNA structure dynamics among different development stages. Also we studied which factors regulate maternal gene decay by RNA structure switch. Overall design: icSHAPE of 12 samples from zebrafish embryos at six different early development stages and 4 samples from Elavl1a deficient zebrafish embryos at sphere and shield stages including two treated with DMSO as control and two treated with NAIN3 for each stage or condition. RNA seq of 6 samples from zebrafish embryos at 2 4 6 h.p.f. and 4 samples from Elavl1a deficient zebrafish embryos at 4 and 6 h.p.f. including two biological replicates. iCLIP seq of 4 samples from flag elavl1a mRNA injected zebrafish embryos at xxx and 6 h.p.f. including two biological replicates.   pubmed:32423473   shield NAIN3 icSHAPE GSM3409387   source name:shield NAIN3|strain:AB|tissue:zebrafish embryos|genotype/variation:normal zebrafish embryos cells|age:6hpf shield NAIN3 icSHAPE Library strategy: icSHAPE icSHAPE computational pipeline reference: PubMedID: 26766114 DOI: 10.1038/nprot.2016.011 trimmed three prime adaptor from reads by cutadapt trimmed low quality bases by trimmomatic collapsed duplicated reads as icSHAPE pipeline trimmed the leading 13nt UMI mapping reads to zebrafish reference transcriptomez10 by bowtie2 using the same parameters in icSHAPE pipeline calculate rpkm as icshape pipeline calculate reverse transcription stop sitesRT stop as icshape pipeline RT stop was normalized by using sliding window then icSHAPE score was calculated as icshape pipeline Genome build: z10 Supplementary files format and content: tab delimited text files for icshape files. First column is the Refseq ID second is transcript length third is RPKM then every column is the nucleotide resolution icSHAPE score. NULL means no confident score. Tab delimited text files for read counts and rpkm files. First column is the Refseq ID second is transcript length third is the unique mapped reads count fourth is the mulitple mapped reads count fifth is the RPKM. shield NAIN3 For in vivo treatment cells was incubated with NAI N3 or DMSO at 28.5 C for 5min. The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol® Reagent invitrogen.To enrich polyA RNA the total RNA was subjected to polyA selection. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37°C for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/µl RiboLock 0.1 U/µl FastAP Life Technology 2 U/µl T4 PNKNEB before 3’ adapter ligation with addition of 10 µl ligation mix 5mM DTT 0.5µM 5’ adenylated and 3’ blocked linker 3’ bio for unmodified 3’ ddc for modified 0.66 U/µl T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25°C for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1µl FastAP 0.2µl SSB Promega 0.8µl Ribolock and 1µl 5’ Deadenylase NEB in 1X NEB buffer 2 NEB at 30°C for 90 min. And then 1µl RecJf NEB was added with another incubation at 37°C for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq， Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer’s protocol. Zebrafish wild type strain AB was raised in system water at 28.5°C under standard conditions. strain:AB|tissue:zebrafish embryos|genotype/variation:normal zebrafish embryos cells|age:6hpf GSM3409387 GSM3409387: shield NAIN3; Danio rerio; OTHER GSM3409387   1 The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol® Reagent invitrogen. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37°C for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/µl RiboLock 0.1 U/µl FastAP Life Technology 2 U/µl T4 PNKNEB before three prime adapter ligation with addition of 10 µl ligation mix 5mM DTT 0.5µM five prime adenylated and three prime blocked linker three prime bio for unmodified three prime ddc for modified 0.66 U/µl T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25°C for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1µl FastAP 0.2µl SSB Promega 0.8µl Ribolock and 1µl five prime Deadenylase NEB in 1X NEB buffer 2 NEB at 30°C for 90 min. And then 1µl RecJf NEB was added with another incubation at 37°C for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq， Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer's protocol. GEO Accession:GSM3409387 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2500   SRP163087     shield_NAIN3_rep1.fastq.gz fastq 152239039350.0 1014926929.0 GSM3409387 r1 0:150 1:0 A:44641737173;C:38963190534;G:30007693087;T:38607452480;N:18966076 150 0     44641737173 38963190534 30007693087 38607452480 18966076 SRX4781863 SRS3862055 SRA787572 GEO Life Science, Tsinghua University 1 0.02906   0.01259   0.97009   0.74209   150   B   usable mapping rate illumina hiseq_era unknown poly_a unknown bulk clip iclip   China 2018-10-01 Gastrula Embryo Embryo Imprecise All anatomical structures
49588 49588 SRR7947912 SRX4781863 SRS3862055 SRP163087 PRJNA494251 Dynamic landscapes and regulome of mRNA structure during zebrafish early embryogenesis GSE120724 Other We resolved the RNA secondary structure during zebrafish early embryogenesis based on in vivo click selective 2' hydroxyl acylation and profiling experiment icSHAPE. We analyzed the RNA structure dynamics among different development stages. Also we studied which factors regulate maternal gene decay by RNA structure switch. Overall design: icSHAPE of 12 samples from zebrafish embryos at six different early development stages and 4 samples from Elavl1a deficient zebrafish embryos at sphere and shield stages including two treated with DMSO as control and two treated with NAIN3 for each stage or condition. RNA seq of 6 samples from zebrafish embryos at 2 4 6 h.p.f. and 4 samples from Elavl1a deficient zebrafish embryos at 4 and 6 h.p.f. including two biological replicates. iCLIP seq of 4 samples from flag elavl1a mRNA injected zebrafish embryos at xxx and 6 h.p.f. including two biological replicates.   pubmed:32423473   shield NAIN3 icSHAPE GSM3409387   source name:shield NAIN3|strain:AB|tissue:zebrafish embryos|genotype/variation:normal zebrafish embryos cells|age:6hpf shield NAIN3 icSHAPE Library strategy: icSHAPE icSHAPE computational pipeline reference: PubMedID: 26766114 DOI: 10.1038/nprot.2016.011 trimmed three prime adaptor from reads by cutadapt trimmed low quality bases by trimmomatic collapsed duplicated reads as icSHAPE pipeline trimmed the leading 13nt UMI mapping reads to zebrafish reference transcriptomez10 by bowtie2 using the same parameters in icSHAPE pipeline calculate rpkm as icshape pipeline calculate reverse transcription stop sitesRT stop as icshape pipeline RT stop was normalized by using sliding window then icSHAPE score was calculated as icshape pipeline Genome build: z10 Supplementary files format and content: tab delimited text files for icshape files. First column is the Refseq ID second is transcript length third is RPKM then every column is the nucleotide resolution icSHAPE score. NULL means no confident score. Tab delimited text files for read counts and rpkm files. First column is the Refseq ID second is transcript length third is the unique mapped reads count fourth is the mulitple mapped reads count fifth is the RPKM. shield NAIN3 For in vivo treatment cells was incubated with NAI N3 or DMSO at 28.5 C for 5min. The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol® Reagent invitrogen.To enrich polyA RNA the total RNA was subjected to polyA selection. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37°C for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/µl RiboLock 0.1 U/µl FastAP Life Technology 2 U/µl T4 PNKNEB before 3’ adapter ligation with addition of 10 µl ligation mix 5mM DTT 0.5µM 5’ adenylated and 3’ blocked linker 3’ bio for unmodified 3’ ddc for modified 0.66 U/µl T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25°C for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1µl FastAP 0.2µl SSB Promega 0.8µl Ribolock and 1µl 5’ Deadenylase NEB in 1X NEB buffer 2 NEB at 30°C for 90 min. And then 1µl RecJf NEB was added with another incubation at 37°C for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq， Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer’s protocol. Zebrafish wild type strain AB was raised in system water at 28.5°C under standard conditions. strain:AB|tissue:zebrafish embryos|genotype/variation:normal zebrafish embryos cells|age:6hpf GSM3409387 GSM3409387: shield NAIN3; Danio rerio; OTHER GSM3409387   1 The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol® Reagent invitrogen. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37°C for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/µl RiboLock 0.1 U/µl FastAP Life Technology 2 U/µl T4 PNKNEB before three prime adapter ligation with addition of 10 µl ligation mix 5mM DTT 0.5µM five prime adenylated and three prime blocked linker three prime bio for unmodified three prime ddc for modified 0.66 U/µl T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25°C for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1µl FastAP 0.2µl SSB Promega 0.8µl Ribolock and 1µl five prime Deadenylase NEB in 1X NEB buffer 2 NEB at 30°C for 90 min. And then 1µl RecJf NEB was added with another incubation at 37°C for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq， Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer's protocol. GEO Accession:GSM3409387 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2500   SRP163087   loader:latf load shield_NAIN3_rep2.fastq.gz fastq 179520653400.0 1196804356.0 GSM3409387 r2 0:150 1:0 A:48366148973;C:44018616729;G:44760216650;T:42350167763;N:25503285 150 0     48366148973 44018616729 44760216650 42350167763 25503285 SRX4781863 SRS3862055 SRA787572 GEO Life Science, Tsinghua University 1 0.03361   0.01615   0.96497   0.718   150   B   usable mapping rate illumina hiseq_era unknown poly_a unknown bulk clip iclip   China 2018-10-01 Gastrula Embryo Embryo Imprecise All anatomical structures
49589 49589 SRR7947909 SRX4781862 SRS3862054 SRP163087 PRJNA494251 Dynamic landscapes and regulome of mRNA structure during zebrafish early embryogenesis GSE120724 Other We resolved the RNA secondary structure during zebrafish early embryogenesis based on in vivo click selective 2' hydroxyl acylation and profiling experiment icSHAPE. We analyzed the RNA structure dynamics among different development stages. Also we studied which factors regulate maternal gene decay by RNA structure switch. Overall design: icSHAPE of 12 samples from zebrafish embryos at six different early development stages and 4 samples from Elavl1a deficient zebrafish embryos at sphere and shield stages including two treated with DMSO as control and two treated with NAIN3 for each stage or condition. RNA seq of 6 samples from zebrafish embryos at 2 4 6 h.p.f. and 4 samples from Elavl1a deficient zebrafish embryos at 4 and 6 h.p.f. including two biological replicates. iCLIP seq of 4 samples from flag elavl1a mRNA injected zebrafish embryos at xxx and 6 h.p.f. including two biological replicates.   pubmed:32423473   shield DMSO icSHAPE GSM3409386   source name:shield DMSO|strain:AB|tissue:zebrafish embryos|genotype/variation:normal zebrafish embryos cells|age:6hpf shield DMSO icSHAPE Library strategy: icSHAPE icSHAPE computational pipeline reference: PubMedID: 26766114 DOI: 10.1038/nprot.2016.011 trimmed three prime adaptor from reads by cutadapt trimmed low quality bases by trimmomatic collapsed duplicated reads as icSHAPE pipeline trimmed the leading 13nt UMI mapping reads to zebrafish reference transcriptomez10 by bowtie2 using the same parameters in icSHAPE pipeline calculate rpkm as icshape pipeline calculate reverse transcription stop sitesRT stop as icshape pipeline RT stop was normalized by using sliding window then icSHAPE score was calculated as icshape pipeline Genome build: z10 Supplementary files format and content: tab delimited text files for icshape files. First column is the Refseq ID second is transcript length third is RPKM then every column is the nucleotide resolution icSHAPE score. NULL means no confident score. Tab delimited text files for read counts and rpkm files. First column is the Refseq ID second is transcript length third is the unique mapped reads count fourth is the mulitple mapped reads count fifth is the RPKM. shield DMSO For in vivo treatment cells was incubated with NAI N3 or DMSO at 28.5 C for 5min. The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol® Reagent invitrogen.To enrich polyA RNA the total RNA was subjected to polyA selection. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37°C for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/µl RiboLock 0.1 U/µl FastAP Life Technology 2 U/µl T4 PNKNEB before 3’ adapter ligation with addition of 10 µl ligation mix 5mM DTT 0.5µM 5’ adenylated and 3’ blocked linker 3’ bio for unmodified 3’ ddc for modified 0.66 U/µl T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25°C for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1µl FastAP 0.2µl SSB Promega 0.8µl Ribolock and 1µl 5’ Deadenylase NEB in 1X NEB buffer 2 NEB at 30°C for 90 min. And then 1µl RecJf NEB was added with another incubation at 37°C for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq， Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer’s protocol. Zebrafish wild type strain AB was raised in system water at 28.5°C under standard conditions. strain:AB|tissue:zebrafish embryos|genotype/variation:normal zebrafish embryos cells|age:6hpf GSM3409386 GSM3409386: shield DMSO; Danio rerio; OTHER GSM3409386   1 The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol® Reagent invitrogen. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37°C for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/µl RiboLock 0.1 U/µl FastAP Life Technology 2 U/µl T4 PNKNEB before three prime adapter ligation with addition of 10 µl ligation mix 5mM DTT 0.5µM five prime adenylated and three prime blocked linker three prime bio for unmodified three prime ddc for modified 0.66 U/µl T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25°C for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1µl FastAP 0.2µl SSB Promega 0.8µl Ribolock and 1µl five prime Deadenylase NEB in 1X NEB buffer 2 NEB at 30°C for 90 min. And then 1µl RecJf NEB was added with another incubation at 37°C for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq， Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer's protocol. GEO Accession:GSM3409386 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2500   SRP163087     shield_DMSO_rep1.fastq.gz fastq 39528129600.0 263520864.0 GSM3409386 r1 0:150 1:0 A:11887790635;C:9698771811;G:8565313614;T:9372993255;N:3260285 150 0     11887790635 9698771811 8565313614 9372993255 3260285 SRX4781862 SRS3862054 SRA787572 GEO Life Science, Tsinghua University 1 0.06286   0.01684   0.93336   0.718   150   B   usable mapping rate illumina hiseq_era unknown poly_a unknown bulk clip iclip   China 2018-10-01 Gastrula Embryo Embryo Imprecise All anatomical structures
49590 49590 SRR7947910 SRX4781862 SRS3862054 SRP163087 PRJNA494251 Dynamic landscapes and regulome of mRNA structure during zebrafish early embryogenesis GSE120724 Other We resolved the RNA secondary structure during zebrafish early embryogenesis based on in vivo click selective 2' hydroxyl acylation and profiling experiment icSHAPE. We analyzed the RNA structure dynamics among different development stages. Also we studied which factors regulate maternal gene decay by RNA structure switch. Overall design: icSHAPE of 12 samples from zebrafish embryos at six different early development stages and 4 samples from Elavl1a deficient zebrafish embryos at sphere and shield stages including two treated with DMSO as control and two treated with NAIN3 for each stage or condition. RNA seq of 6 samples from zebrafish embryos at 2 4 6 h.p.f. and 4 samples from Elavl1a deficient zebrafish embryos at 4 and 6 h.p.f. including two biological replicates. iCLIP seq of 4 samples from flag elavl1a mRNA injected zebrafish embryos at xxx and 6 h.p.f. including two biological replicates.   pubmed:32423473   shield DMSO icSHAPE GSM3409386   source name:shield DMSO|strain:AB|tissue:zebrafish embryos|genotype/variation:normal zebrafish embryos cells|age:6hpf shield DMSO icSHAPE Library strategy: icSHAPE icSHAPE computational pipeline reference: PubMedID: 26766114 DOI: 10.1038/nprot.2016.011 trimmed three prime adaptor from reads by cutadapt trimmed low quality bases by trimmomatic collapsed duplicated reads as icSHAPE pipeline trimmed the leading 13nt UMI mapping reads to zebrafish reference transcriptomez10 by bowtie2 using the same parameters in icSHAPE pipeline calculate rpkm as icshape pipeline calculate reverse transcription stop sitesRT stop as icshape pipeline RT stop was normalized by using sliding window then icSHAPE score was calculated as icshape pipeline Genome build: z10 Supplementary files format and content: tab delimited text files for icshape files. First column is the Refseq ID second is transcript length third is RPKM then every column is the nucleotide resolution icSHAPE score. NULL means no confident score. Tab delimited text files for read counts and rpkm files. First column is the Refseq ID second is transcript length third is the unique mapped reads count fourth is the mulitple mapped reads count fifth is the RPKM. shield DMSO For in vivo treatment cells was incubated with NAI N3 or DMSO at 28.5 C for 5min. The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol® Reagent invitrogen.To enrich polyA RNA the total RNA was subjected to polyA selection. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37°C for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/µl RiboLock 0.1 U/µl FastAP Life Technology 2 U/µl T4 PNKNEB before 3’ adapter ligation with addition of 10 µl ligation mix 5mM DTT 0.5µM 5’ adenylated and 3’ blocked linker 3’ bio for unmodified 3’ ddc for modified 0.66 U/µl T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25°C for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1µl FastAP 0.2µl SSB Promega 0.8µl Ribolock and 1µl 5’ Deadenylase NEB in 1X NEB buffer 2 NEB at 30°C for 90 min. And then 1µl RecJf NEB was added with another incubation at 37°C for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq， Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer’s protocol. Zebrafish wild type strain AB was raised in system water at 28.5°C under standard conditions. strain:AB|tissue:zebrafish embryos|genotype/variation:normal zebrafish embryos cells|age:6hpf GSM3409386 GSM3409386: shield DMSO; Danio rerio; OTHER GSM3409386   1 The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol® Reagent invitrogen. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37°C for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/µl RiboLock 0.1 U/µl FastAP Life Technology 2 U/µl T4 PNKNEB before three prime adapter ligation with addition of 10 µl ligation mix 5mM DTT 0.5µM five prime adenylated and three prime blocked linker three prime bio for unmodified three prime ddc for modified 0.66 U/µl T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25°C for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1µl FastAP 0.2µl SSB Promega 0.8µl Ribolock and 1µl five prime Deadenylase NEB in 1X NEB buffer 2 NEB at 30°C for 90 min. And then 1µl RecJf NEB was added with another incubation at 37°C for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq， Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer's protocol. GEO Accession:GSM3409386 OTHER TRANSCRIPTOMIC other SINGLE ILLUMINA Illumina HiSeq 2500   SRP163087     shield_DMSO_rep2.fastq.gz fastq 43632084450.0 290880563.0 GSM3409386 r2 0:150 1:0 A:12666468004;C:10421063446;G:9540808484;T:10977192542;N:26551974 150 0     12666468004 10421063446 9540808484 10977192542 26551974 SRX4781862 SRS3862054 SRA787572 GEO Life Science, Tsinghua University 1 0.08049   0.02275   0.91616   0.64879   150   B   usable mapping rate illumina hiseq_era unknown poly_a unknown bulk clip iclip   China 2018-10-01 Gastrula Embryo Embryo Imprecise All anatomical structures
50540 50540 SRR8134463 SRX4955489 SRS3996626 SRP167225 PRJNA501843 Characterization of Transcriptomic Profile in Early Zebrafish PGCs by Single Cell Sequencing PRJNA501843 Other Single cell RNA seq was applied for studying the transcriptomic profile in early zebrafish PGCsprimordial germ cells by choosing three time points during zebrafish embryonic development. The three time points were 6hpfhpf also called shield stage 11hpfalso called 3 somite stage and 24hpfalso called prim 5 stage.         H6 3   strain:AB|isolate:missing|breed:missing|cultivar:missing|ecotype:missing|dev stage:6hpf biological replicate 3|sex:missing|tissue:PGC|BioSampleModel:Model organism or animal                 RNA Seq of Danio rerio: PGC H6 3 H6 3 smart2     RNA-Seq TRANSCRIPTOMIC other PAIRED ILLUMINA Illumina HiSeq 2500   SRP167225     H6_3_R1.fastq H6_3_R2.fastq fastq fastq 989016500.0 3956066.0 H6 3 R1.fastq 0:125 1:125 A:270941686;C:224568911;G:226264637;T:267238750;N:2516 125 125     270941686 224568911 226264637 267238750 2516 SRX4955489 SRS3996626 SRA800727 Shanghai Institute of Biochemistry and Cell Biology, CAS|State Key Laboratory of cell Biology Shanghai Institute of Biochemistry and Cell Biology, CAS 2 0.94889 0.94713 0.02757 0.02789 0.84415 0.84571 0.52218 0.52291 125 125 B B biological fallback assumption illumina hiseq_era unknown other unknown sc_generic single_cell_generic generic-scrnaseq-only   China 2019-02-01 Gastrula Embryo Undetermined Embryo Imprecise
50542 50542 SRR8134465 SRX4955487 SRS3996624 SRP167225 PRJNA501843 Characterization of Transcriptomic Profile in Early Zebrafish PGCs by Single Cell Sequencing PRJNA501843 Other Single cell RNA seq was applied for studying the transcriptomic profile in early zebrafish PGCsprimordial germ cells by choosing three time points during zebrafish embryonic development. The three time points were 6hpfhpf also called shield stage 11hpfalso called 3 somite stage and 24hpfalso called prim 5 stage.         H6 1   strain:AB|isolate:missing|breed:missing|cultivar:missing|ecotype:missing|dev stage:6hpf biological replicate 1|sex:missing|tissue:PGC|BioSampleModel:Model organism or animal                 RNA Seq of Danio rerio: PGC H6 1 H6 1 smart2     RNA-Seq TRANSCRIPTOMIC other PAIRED ILLUMINA Illumina HiSeq 2500   SRP167225     H6_1_R1.fastq H6_1_R2.fastq fastq fastq 759809500.0 3039238.0 H6 1 R1.fastq 0:125 1:125 A:211499426;C:170044275;G:171920878;T:206342561;N:2360 125 125     211499426 170044275 171920878 206342561 2360 SRX4955487 SRS3996624 SRA800727 Shanghai Institute of Biochemistry and Cell Biology, CAS|State Key Laboratory of cell Biology Shanghai Institute of Biochemistry and Cell Biology, CAS 2 0.94965 0.94791 0.04309 0.04276 0.83485 0.83676 0.5519 0.56156 125 125 B B biological fallback assumption illumina hiseq_era unknown other unknown sc_generic single_cell_generic generic-scrnaseq-only   China 2019-02-01 Gastrula Embryo Undetermined Embryo Imprecise
50543 50543 SRR8134466 SRX4955486 SRS3996625 SRP167225 PRJNA501843 Characterization of Transcriptomic Profile in Early Zebrafish PGCs by Single Cell Sequencing PRJNA501843 Other Single cell RNA seq was applied for studying the transcriptomic profile in early zebrafish PGCsprimordial germ cells by choosing three time points during zebrafish embryonic development. The three time points were 6hpfhpf also called shield stage 11hpfalso called 3 somite stage and 24hpfalso called prim 5 stage.         H6 2   strain:AB|isolate:missing|breed:missing|cultivar:missing|ecotype:missing|dev stage:6hpf biological replicate 2|sex:missing|tissue:PGC|BioSampleModel:Model organism or animal                 RNA Seq of Danio rerio: PGC H6 2 H6 2 smart2     RNA-Seq TRANSCRIPTOMIC other PAIRED ILLUMINA Illumina HiSeq 2500   SRP167225     H6_2_R1.fastq H6_2_R2.fastq fastq fastq 858125000.0 3432500.0 H6 2 R1.fastq 0:125 1:125 A:236752508;C:193146437;G:194455657;T:233768197;N:2201 125 125     236752508 193146437 194455657 233768197 2201 SRX4955486 SRS3996625 SRA800727 Shanghai Institute of Biochemistry and Cell Biology, CAS|State Key Laboratory of cell Biology Shanghai Institute of Biochemistry and Cell Biology, CAS 2 0.94359 0.93966 0.04047 0.03943 0.83924 0.84396 0.54153 0.54602 125 125 B B biological fallback assumption illumina hiseq_era unknown other unknown sc_generic single_cell_generic generic-scrnaseq-only   China 2019-02-01 Gastrula Embryo Undetermined Embryo Imprecise
56343 56343 SRR10948889 SRX7615947 SRS6049205 SRP243873 PRJNA602610 Global promoter usage in the differentfractions of the cell during early zebrafish development GSE144040 Other We employed transcriptomics methods to examine differences in promoter usage in the nuclear and cytosolic fractions of zebrafish embryonic cells at different stages of development. The analysis of the CAGE seq data revealed differences in promoter width within the same stage Post MZT in different compartments of the embryonic cell proposing a spatial and temporal regulation of gene expression. Overall design: Promoter organisation in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome Shield and Prim 5 stages of development.   pubmed:33795334   Shield Cyt CAGE GSM4278495   source name:Shield stage embryo|tissue:zebrafish embryos|Stage:Shield|fraction:Cytosolic|molecule type:capped RNAs Shield Cyt CAGE Library strategy: LQ CAGE sequencing The raw tags from CAGE sequencing were mapped using STAR aligner and the resulting BAM files were used in the bioconductor package CAGEr for further downstream analysis. CAGEr starts from mapped reads and does quality filtering normalization removal of the additional 5’ end G nucleotide added during the CAGE protocol and the frequency of the usage of start sites Haberle et al. 2015. Genome build: Zv9 Supplementary files format and content: CTSS files in bedGraph format Shield stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit The conventional method used for 5’ cap analysis also called cap trapping method Carninci et al. 1996 targets all capped mRNAs by chemical modification of the 5’ end cap followed by its biotinylation and then a pulldown using streptavidin coated beads. We used a modified version of cap trapping called Low Quantity CAGE or LQ CAGE for our CAGE library preparation. This method allowed use of limited quantities of RNAs as is generally obtained in early stages of development. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|Stage:Shield|fraction:Cytosolic|molecule type:capped RNAs GSM4278495 GSM4278495: Shield Cyt CAGE; Danio rerio; OTHER GSM4278495   1 RNA was extracted using the Rneasy mini kit The conventional method used for five prime cap analysis also called cap trapping method Carninci et al. 1996 targets all capped mRNAs by chemical modification of the five prime end cap followed by its biotinylation and then a pulldown using streptavidin coated beads. We used a modified version of cap trapping called Low Quantity CAGE or LQ CAGE for our CAGE library preparation. This method allowed use of limited quantities of RNAs as is generally obtained in early stages of development. GEO Accession:GSM4278495 OTHER TRANSCRIPTOMIC other PAIRED ILLUMINA Illumina HiSeq 2500   SRP243873     Shield_Cyt_L006_R1_001.fastq.gz Shield_Cyt_L006_R2_001.fastq.gz fastq fastq 8704643185.0 91627823.0 GSM4278495 r1 0:51 1:44 A:1930221949;C:2128885351;G:2388442525;T:2256907064;N:186296 51 44     1930221949 2128885351 2388442525 2256907064 186296 SRX7615947 SRS6049205 SRA1029879 GEO School of Biosciences, University of Birmingham 2 0.89389 0.40438 0.14385 0.06093 0.78792 0.8352 0.5381 0.52296 51 44 B B mate2-mate1 similar by mapping diff illumina hiseq_era 5prime other unknown bulk unknown unknown   United Kingdom 2020-01-22 Gastrula Embryo Embryo Imprecise All anatomical structures
56344 56344 SRR10948888 SRX7615946 SRS6049206 SRP243873 PRJNA602610 Global promoter usage in the differentfractions of the cell during early zebrafish development GSE144040 Other We employed transcriptomics methods to examine differences in promoter usage in the nuclear and cytosolic fractions of zebrafish embryonic cells at different stages of development. The analysis of the CAGE seq data revealed differences in promoter width within the same stage Post MZT in different compartments of the embryonic cell proposing a spatial and temporal regulation of gene expression. Overall design: Promoter organisation in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome Shield and Prim 5 stages of development.   pubmed:33795334   Shield Nuc CAGE GSM4278494   source name:Shield stage embryo|tissue:zebrafish embryos|Stage:Shield|fraction:Nuclear|molecule type:capped RNAs Shield Nuc CAGE Library strategy: LQ CAGE sequencing The raw tags from CAGE sequencing were mapped using STAR aligner and the resulting BAM files were used in the bioconductor package CAGEr for further downstream analysis. CAGEr starts from mapped reads and does quality filtering normalization removal of the additional 5’ end G nucleotide added during the CAGE protocol and the frequency of the usage of start sites Haberle et al. 2015. Genome build: Zv9 Supplementary files format and content: CTSS files in bedGraph format Shield stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit The conventional method used for 5’ cap analysis also called cap trapping method Carninci et al. 1996 targets all capped mRNAs by chemical modification of the 5’ end cap followed by its biotinylation and then a pulldown using streptavidin coated beads. We used a modified version of cap trapping called Low Quantity CAGE or LQ CAGE for our CAGE library preparation. This method allowed use of limited quantities of RNAs as is generally obtained in early stages of development. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|Stage:Shield|fraction:Nuclear|molecule type:capped RNAs GSM4278494 GSM4278494: Shield Nuc CAGE; Danio rerio; OTHER GSM4278494   1 RNA was extracted using the Rneasy mini kit The conventional method used for five prime cap analysis also called cap trapping method Carninci et al. 1996 targets all capped mRNAs by chemical modification of the five prime end cap followed by its biotinylation and then a pulldown using streptavidin coated beads. We used a modified version of cap trapping called Low Quantity CAGE or LQ CAGE for our CAGE library preparation. This method allowed use of limited quantities of RNAs as is generally obtained in early stages of development. GEO Accession:GSM4278494 OTHER TRANSCRIPTOMIC other PAIRED ILLUMINA Illumina HiSeq 2500   SRP243873     Shield_Nuc_L006_R1_001.fastq.gz Shield_Nuc_L006_R2_001.fastq.gz fastq fastq 1700102330.0 17895814.0 GSM4278494 r1 0:51 1:44 A:370009933;C:422952763;G:478245997;T:428857334;N:36303 51 44     370009933 422952763 478245997 428857334 36303 SRX7615946 SRS6049206 SRA1029879 GEO School of Biosciences, University of Birmingham 2 0.9163 0.35706 0.26264 0.1183 0.78244 0.863 0.6044 0.56423 51 44 B T mate2 technical by mapping diff illumina hiseq_era 5prime other unknown bulk unknown unknown   United Kingdom 2020-01-22 Gastrula Embryo Embryo Imprecise All anatomical structures
72833 72833 SRR23210472 SRX19158411 SRS16570832 SRP418915 PRJNA927004 Differential Nodal level promotes mesendoderm cell fate segregation mediated by chromatin organization GSE223636 Other Purpose: To investigate the mechanism of prechordal plate and anterior endoderm separation . Methods: Nodal injected explants injected with 10pg ndr2 mRNA constructed from lft1 mutants and ndr1 morphants were harvested at xxxhpf. Libraries were prepared using Chromium Controller and Chromium Single Cell three primeLibrary & Gel Bead Kit v3 10x Genomics PN 1000075 according to the manufacturer's protocol for 10000 cells recovery. For single cell multiomics zebrafish embryos at 6 hpf were harvested. Libraries were prepared using Chromium Next GEM Single Cell Multiome ATAC + Gene Expression Reagent Bundle 10x Genomics 4 rxns PN 1000285 according to the manufacturer's protocol for 10000 cells recovery. Results: A total of 10 614 single cell transcriptomes and 4 335 multiomics were collected post stringent quality control measures. Conclusions: A slight bias in Nodal signaling promotes a differential chromatin structure between prechordal plate and endoderm which drives a differential expression of those key regulators such as gsc and ripply1 in these two cell lineages and further regulates mesendoderm cell fate separation. Overall design: zebrafish Nodal explants constructed from lft1 mutants and ndr1 morphants were harvested at 6hpf for scRNA seq. Zebrafish embryos were harvested at 6hpf for single cell multiomics.       zebrafish embryo multiomics expression GSM6969677   source name:zebrafish cells|strain:AB|tissue:embryonic cells|age:6hpf zebrafish embryo multiomics expression Illumina sequencing reads were aligned to the zebrafish mRNA reference genome GRCz11 using the 10x Genomics CellRanger pipeline version 6.1.2 and cellranger arc version 2.0.0 with default parameters. Assembly: GRCz11 Supplementary files format and content: tar archivr or gzip compressed files included filtered gene bc matrices and ATAC fragments post running CellRanger or cellranger arc pipeline. Library strategy: scMultiome GEX zebrafish cells 10pg of ndr2 mRNA was injected to one cell of embryonic animal pole at xxx cell stage. All embryos were incubated in 0.3x Danieau buffer until 1k stage then were transferred to Dulbecco's Modified Eagle Medium. Animal pole explants corresponding roughly to half of the blastula were incubated to 6hpf corresponding to embryonic developmental stage. Libraries were prepared using Chromium Controller and Chromium Single Cell 3’Library & Gel Bead Kit v3 10x Genomics PN 1000075 and Chromium Next GEM Single Cell Multiome ATAC + Gene Expression Reagent Bundle 10x Genomics 4 rxns PN 1000285 according to the manufacturer’s protocol for 10000 cells recovery. Explants were cultured in a Petri dish coated with 1.5% agarose filled with Dulbecco's Modified Eagle Medium strain:AB|tissue:embryonic cells|age:6hpf GSM6969677 GSM6969677: zebrafish embryo multiomics expression; Danio rerio; OTHER GSM6969677 r1 GSM6969677 1 Libraries were prepared using Chromium Controller and Chromium Single Cell three primeLibrary & Gel Bead Kit v3 10x Genomics PN 1000075 and Chromium Next GEM Single Cell Multiome ATAC + Gene Expression Reagent Bundle 10x Genomics 4 rxns PN 1000285 according to the manufacturer's protocol for 10000 cells recovery.   OTHER TRANSCRIPTOMIC other PAIRED ILLUMINA Illumina NovaSeq 6000   SRP418915   loader:fastq load.py 6-M-GEX-merge_S1_L001_R1_001.fastq.gz 6-M-GEX-merge_S1_L001_R2_001.fastq.gz fastq fastq 67059456600.0 223531522.0 GSM6969677 r1 0:150 1:150 A:20681044309;C:12370348813;G:11664316954;T:22341885756;N:1860768 150 150     20681044309 12370348813 11664316954 22341885756 1860768 SRX19158411 SRS16570832 SRA1581154 Institute of genetics, Zhejiang University Institute of genetics, Zhejiang University 2 0.45314 0.8745 0.16489 0.2261 0.99072 0.82641 0.79221 0.75799 150 150 T B mate1 technical by mapping diff illumina novaseq_era unknown other unknown sc single_cell_droplet 10x   China 2023-01-24 Gastrula Embryo Embryo Imprecise All anatomical structures

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CREATE TABLE run_metadata("run.accession" VARCHAR, "experiment.accession" VARCHAR, "sample.accession" VARCHAR, "study.accession" VARCHAR, bioproject VARCHAR, "study.title" VARCHAR, "study.alias" VARCHAR, "study.type" VARCHAR, "study.abstract" VARCHAR, "study.attributes" VARCHAR, "study.PMIDs" VARCHAR, "sample.description" VARCHAR, "sample.title" VARCHAR, "sample.alias" VARCHAR, "sample.centername" VARCHAR, "sample.attributes" VARCHAR, "GEOsample.title" VARCHAR, "GEOsample.dataprocessing" VARCHAR, "GEOsample.source" VARCHAR, "GEOsample.treatmentprotocol" VARCHAR, "GEOsample.extractprotocol" VARCHAR, "GEOsample.growthprotocol" VARCHAR, "GEOsample.characteristics" VARCHAR, "GEOsample.accession" VARCHAR, "experiment.title" VARCHAR, "experiment.alias" VARCHAR, "experiment.library_name" VARCHAR, "experiment.design_description" VARCHAR, "experiment.library_construction_protocol" VARCHAR, "experiment.attributes" VARCHAR, "experiment.library_strategy" VARCHAR, "experiment.library_source" VARCHAR, "experiment.library_selection" VARCHAR, "experiment.library_layout" VARCHAR, "experiment.platform" VARCHAR, "experiment.instrument_model" VARCHAR, "experiment.spot_descriptor" VARCHAR, "experiment.study_ref" VARCHAR, "run.title" VARCHAR, "run.attributes" VARCHAR, "run.filename" VARCHAR, "run.semantic_name" VARCHAR, "run.total_bases" DOUBLE, "run.total_spots" DOUBLE, "run.alias" VARCHAR, "run.read_lengths" VARCHAR, "run.base_counts" VARCHAR, "run.r1_length" BIGINT, "run.r2_length" BIGINT, "run.r3_length" BIGINT, "run.r4_length" BIGINT, "run.Acount" BIGINT, "run.Ccount" BIGINT, "run.Gcount" BIGINT, "run.Tcount" BIGINT, "run.Ncount" BIGINT, "run.experiment" VARCHAR, "run.pool_member" VARCHAR, "submission.accession" VARCHAR, "submission.srasource" VARCHAR, "submission.bioprojectsource" VARCHAR, "seqdetective.n_mates" BIGINT, "seqdetective.mapping_rate.mate1" DOUBLE, "seqdetective.mapping_rate.mate2" DOUBLE, "seqdetective.nofeature_rate.mate1" DOUBLE, "seqdetective.nofeature_rate.mate2" DOUBLE, "seqdetective.sparsity.mate1" DOUBLE, "seqdetective.sparsity.mate2" DOUBLE, "seqdetective.pos_strand_rate.mate1" DOUBLE, "seqdetective.pos_strand_rate.mate2" DOUBLE, "seqdetective.readlen.mate1" BIGINT, "seqdetective.readlen.mate2" BIGINT, "seqdetective.judgement.mate1" VARCHAR, "seqdetective.judgement.mate2" VARCHAR, "seqdetective.judgement.reason" VARCHAR, platform_family VARCHAR, instrument_generation VARCHAR, read_bias VARCHAR, selection_class VARCHAR, prep_kit VARCHAR, sc_or_bulk VARCHAR, tech_class VARCHAR, technology VARCHAR, tech_variant VARCHAR, "submission.bioprojectsource.country" VARCHAR, earliest_date DATE, devstage_curation VARCHAR, devstage_curation_coarse VARCHAR, tissue_curation VARCHAR, tissue_curation_coarse VARCHAR);;
CREATE INDEX idx_run_bioproject ON run_metadata(bioproject);;
CREATE INDEX idx_run_run_accession ON run_metadata("run.accession");;