{"database": "metadata", "table": "run_metadata", "rows": [[37120, "SRR942766", "SRX327592", "SRS463964", "SRP028229", "PRJNA213244", "Autophagy regulator DRAM1 functions downstream of MYD88 in defense against tuberculosis RNA seq", "GSE49186", "Transcriptome Analysis", "MyD88 is an adaptor protein in Toll like receptor and interleukin 1 receptor mediated signaling pathways that plays an essential role in activation of immune responses following pathogen recognition. We investigate that role in the zebrafish embryo model by using a zebrafish mutant line that contains a premature stop condon in the gene encoding MyD88  leading to a truncated protein that lacks domains important for its normal function. We infected these MyD88 mutants and wildtype individuals with Mycobacterium marinum to compare the resulting immune response by transcriptome profiling on total RNA isolated from single embryos. Autophagy regulator dram1 was identified as one of the MyD88 dependent genes. Overall design: This RNAseq analysis was used to determine the effect of a truncation of the MyD88 protein on the innate immune response of zebrafish embryos during infection with Mycobacterium marinum. Myd88 mutant and wild type embryos were derived by incrossing homozygous myd88 mutant parents allele hu3568  van der Vaart et al.  2013  Disease models & mechanisms 6  841 854 or their wildtype siblings. RNA was isolated from pools of 20 embryos at 4 days post infection 4 dpi.  The following treatment groups were used: homozygous mutants mock injected with PBS/2%PVP 4 dpi  2 wildtype siblings mock injected with PBS/2%PVP 4dpi  3 M. marinum infected homozygous mutants 4dpi  4 M. marinum infected wildtype siblings 4dpi. Embryos were grown at 28.5\u201330\u00b0C in egg water and manually dechorionated at 24 hpf. Subsequently  embryos were infected at 28 hpf by micro injecting 200 colony forming units CFU of Mycobacterium marinum Mma20 bacteria into the caudal vein  or were mock injected with buffer PBS/2%PVP as a control.  post injections embryos were transferred into fresh egg water and incubated for 4 days at 28\u00b0C. post the incubation period  single embryos were snap frozen in liquid nitrogen and RNA was isolated for RNAseq analysis.", "parent bioproject:PRJNA213243", "pubmed:24922577;pubmed:25247677", null, "myd88 /  uninfected", "GSM1195170", null, "source name:myd88 mutant myd88 /   not infected|sample type:myd88 mutant zebrafish embryos mock injected with PBS at xxx hpf; RNA isolated at 4 dpi|treatment:PBS|genotype:myd88 mutant|tissue:embryo", "myd88 /  uninfected", "Image analysis and base calling was done by the Illumina HCS version 1.15.1; raw data from 2 sample 20 and 3 sequencing runs samples 19  21  and 22 were combined  to generate the tsv files and to calculate average insert sizes and standard deviations for paired end experiments. Sequence reads were quality trimmed using the quality trim module in the CLCbio Assembly Cell v4.0.6. Filtered reads were mapped to Ensembl transcripts Zv9 67 using the ref assemle short module in the CLCbio Assembly Cell v4.0.6. Accumulation of transcripts to Ensembl genes was done by first converting the mapping files to a table with the assembly table module in the CLCbio Assembly Cell v4.0.6. Secondly  a custom script was used that sums all reads belonging to the same gene. Non uniquely mapped reads were divided between genes according to their ratio of uniquely mapped reads. Finally  read counts of transcripts belonging to the same gene were summed to obtain count data at Ensembl gene level. Fold change and differential expression significance values were calculated from gene level read counts using the DESeq package version 1.8.3 available in Bioconductor version 2.10. Genome build: Danio rerio.Zv9.67.dna.toplevel / Danio rerio.Zv9.67.cdna.all.fa http://www.ensembl.org/info/data/ftp/index.html  Supplementary files format and content: tab delimited text files include RPKM values for each Sample", "myd88 mutant myd88 /   not infected", "Zebrafish embryos were manually dechorionated at 24 hpf and at 28 hpf they were micro injected into the caudal vein with 200 CFU of M. marinum Mma20 bacteria suspended in PBS/2%PVP or mock injected with PBS/2%PVP as a control. At 5 dpf 4 d post infection pools of 20 embryos per treatment group were snap frozen in liquid nitrogen  and total RNA was isolated using TRIZOL reagent.", "Pools of embryos for RNA isolation were snap frozen in liquid nitrogen and subsequently stored at  80\u00b0C. Embryos were homogenized in 1 ml of TRIZOL\u00ae Reagent Invitrogen and subsequently total RNA was extracted according to the manufacturer\u2019s instructions. The RNA samples were incubated for 20 min at 37\u00b0 with 10 units of DNaseI Roche Applied Science to remove residual genomic DNA prior to purification using the RNeasy MinElute Cleanup kit Qiagen according to the RNA clean up protocol. The integrity of the RNA was confirmed by Lab on chip analysis using the 2100 Bioanalyzer Agilent Technologies. A total of 3 \u03bcg of RNA was used to make RNAseq libraries using the Illumina TruSeq RNA Sample Preparation Kit v2 Illumina Inc.  San Diego  USA. In the manufacturer\u2019s instructions two modifications were made. In the adapter ligation step 1 \u00b5l instead of 2.5 \u00b5l adapter was used. In the library size selection step the library fragments were isolated with a double Ampure XP purification with a 0.7x beads to library ratio. The resulting mRNA Seq library was sequenced using an Illumina HiSeq2000 instrument according to the manufacturer\u2019s description with a read length of 2 x 50 nucleotides.", "Zebrafish were handled in compliance with the local animal welfare regulations and maintained according to standard protocols http://ZFIN.org. Embryos were grown at 28 5\u00b0C in egg water 60\u00b5g/ml Instant Ocean sea salts.", "sample type:myd88 mutant zebrafish embryos mock injected with PBS at xxx hpf; RNA isolated at 4 dpi|treatment:PBS|genotype:myd88 mutant|tissue:embryo", "GSM1195170", "GSM1195170: myd88 /  uninfected; Danio rerio; RNA Seq", "GSM1195170", null, "1", "Pools of embryos for RNA isolation were snap frozen in liquid nitrogen and subsequently stored at  80\u00b0C. Embryos were homogenized in 1 ml of TRIZOL\u00ae Reagent Invitrogen and subsequently total RNA was extracted according to the manufacturer\u2019s instructions. The RNA samples were incubated for 20 min at 37\u00b0 with 10 units of DNaseI Roche Applied Science to remove residual genomic DNA prior to purification using the RNeasy MinElute Cleanup kit Qiagen according to the RNA clean up protocol. The integrity of the RNA was confirmed by Lab on chip analysis using the 2100 Bioanalyzer Agilent Technologies. A total of 3 \u03bcg of RNA was used to make RNAseq libraries using the Illumina TruSeq RNA Sample Preparation Kit v2 Illumina Inc.  San Diego  USA. In the manufacturer\u2019s instructions two modifications were made. In the adapter ligation step 1 \u00b5l instead of 2.5 \u00b5l adapter was used. In the library size selection step the library fragments were isolated with a double Ampure XP purification with a 0.7x beads to library ratio. The resulting mRNA Seq library was sequenced using an Illumina HiSeq2000 instrument according to the manufacturer\u2019s description with a read length of 2 x 50 nucleotides.", "GEO Accession:GSM1195170", "RNA-Seq", "TRANSCRIPTOMIC", "cDNA", "PAIRED", "ILLUMINA", "Illumina HiSeq 2000", null, "SRP028229", null, null, "19_AGTTCC_L008_R1_001_run3.fastq.gz 19_AGTTCC_L008_R2_001_run3.fastq.gz", "fastq fastq", 73803018.0, 723559.0, "GSM1195170 r3", "0:51 1:51", "A:19853585;C:17320732;G:16864909;T:19733714;N:30078", 51, 51, null, null, 19853585, 17320732, 16864909, 19733714, 30078, "SRX327592", "SRS463964", "SRA096049", "GEO", "Animal Sciences and Health, Institute of Biology, Leiden University", 2, 0.92306, 0.91503, 0.11783, 0.1182, 0.67381, 0.67389, 0.4768, 0.47693, 51, 51, "B", "B", "biological fallback assumption", "illumina", "hiseq_era", "unknown", "poly_a", "trueseq", "bulk", "unknown", "unknown", null, "Netherlands", "2013-07-24", "Multi-stage", "Multi-stage", "Embryo Imprecise", "All anatomical structures"]], "columns": ["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"], "primary_keys": ["rowid"], "primary_key_values": ["37120"], "units": {}, "query_ms": 9.230715004377998}