run_metadata: 65567

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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
65567	SRR15373448	SRX11676000	SRS9707982	SRP331635	PRJNA753019	five prime half of specific tRNAs feeds back to promote corresponding tRNA gene transcription in vertebrate embryos [tRNA seq]	GSE181683	Transcriptome Analysis	five primetRFls are small tRNA fragments derived from five prime half of mature tRNAs. However it is unknown whether five primetRFls could feed back to regulate tRNA biogenesis. Here we show that five primetRFlGly/GCC and five primetRFlGlu/CTC function to promote transcription of corresponding tRNA genes and are essential for vertebrate early embryogenesis. During zebrafish embryogenesis dynamics of five primetRFlGly/GCC and five primetRFlGlu/CTC levels correlates with that of tRNAGly/GCC and tRNAGlu/CTC levels. Morpholino mediated knockdown of five primetRFlGly/GCC or five primetRFlGlu/CTC down regulates tRNAGly/GCC or tRNAGlu/CTC levels respectively and causes embryonic lethality that is efficiently rescued by co injection of properly refolded corresponding tRNA. In zebrafish embryos tRNA:DNA and five primetRFl:DNA hybrids commonly exist on the template strand of tRNA genes. Mechanistically unstable five primetRFl:DNA hybrid may prevent the formation of transcriptionally inhibitory stable tRNA:DNA hybrids on the same tRNA loci so as to facilitate tRNA genes transcription. The uncovered mechanism may be implicated in other physiological and pathological processes. Overall design: tRNAs from wildtype zebrafish eggs and early embryos were detected by high throughput sequencing. Total RNAs from mature egg 1cell 256c sphere shield and 24hpf stage embryos were extracted by Trizol reagent. Then 60 120 nt length RNA was enriched by separation and elution from denature gel. RNA was treated with a combination of ALKB protein to remove some modifications before library preparation.	parent bioproject:PRJNA753013	pubmed:34797706		tRNA 1cell	GSM5509227		tissue:Whole Embryo\|genotype:WT\|strain:TU\|developmental stage:1cell	tRNA 1cell	Adaptor trimming and quality control were done by FastP Version 0.19.5 and Cutadapt Version 3.4 then 60 100 nt length reads were preserved for down stream analysis. Reads were first mapped to genome GRCz11 by Bowtie2 Version 2.4.2 with very sensitive local mode D 20 R 3 N 0 L 20 i S 1 0.50. Then aligned reads were further mapped to tRNA database GtRNAdb GRCz11. The counts of reads aligned to each tRNAs were calculated and normalized to all reads mapped to genome for down stream analysis. Genome build: GRCz11 Genome build: tRNA database from GtRNAdb GRCz11 Supplementary files format and content: tab delimited text files include tRNA RPM values for each sample	Whole Embryo		About 50 100 zebrafish eggs or embryos at a desired stage were collected to extract total RNA and dechorionated by Pronase digestion. Embryos were then transferred to 1.5 ml eppendorf tube and lysed in 1 ml Trizol Reagent Thermo Fisher Scientific 15596018. The lysate was centrifuged at 12 000 rpm for 10 min and the supernatant was collected and mixed with 200 μl chloroform in a fresh tube. post centrifugation at 12 000 rpm for 10 min at 4℃ the supernatant was transferred to a fresh tube with addition of equal volume of isopropanol and incubated at room temperature for 15 min followed by thorough mix and centrifugation at 12 000 rpm for 10 20 min. The RNA pellet was washed by 75% ethanol once and was stored as pellet in 75% ethanol at 80℃ until all samples were collected. Total RNA was placed on ice for 30 min with 10% PEG8000 and 0.5 M NaCl then centrifuged at 14 000 rpm for 15 min at 4℃. Supernatant low molecular weight LMW RNA was collected and precipitated by addition of ethanol. The LMW RNA was separated by 8% 8 M Urea PAGE gel then the bands correspond to 60 120 nt length were cut and crunched into small pieces. RNA was eluted from the gel by 0.3 M NaOAc overnight on a vertical rotator at 4℃. Then the supernatant was precipitated at 20℃ post addition with 2 μl of Glycoblue and 2.5 volume of ethanol. post centrifuged at 12 000 rpm for 30 min and washed by 75% ethanol the RNA precipitant was dissolved in RNase free water. There are three pretreatment steps before library preparation. 60 120nt RNA enriched in the previous step was first incubated in 100 mM Tris HCl pH 9.0 at 37℃ for 30 min to be deacylated then the RNA was treated with T4 PNK to repair ends at 37℃ for 30 min. Modifications on tRNAs were removed by ALKB treatment [1 μg purified RNA was treated by ALKB protein mixture wt ALKB 160 pmol; ALKB D135S 200 pmol; and ALKB D135S/L118V 200 pmol in reaction buffer 300 mM KCl 2 mM MgCl2 50 μM NH42FeSO42 300 μM α ketoglutarate 2 mM vitamin C 50 μg/ml BSA 50 mM MES pH 5.0 40 U RNase Inhibitor 1x Protease Inhibitor for 2 h at room temperature]. Purification was needed post each pre treatment step. 100 ng RNA was then subjected for RNA library preparation. tRNA library was prepared by VAHTS Small RNA Library Prep Kit for Illumina Vazyme NR801. RNA was heat denatured at 80℃ for 2 min in the first step then every steps were done followed the manual in kit. The libraries were first concentrated by DNA concentrator spin column Zymo then separated on 6% native PAGE gel. Bands correspond to 180 240 bp 60 120 nt RNA ligated with both adaptors length were cut and DNA libraries were eluted from gels.		genotype:WT\|strain:TU\|developmental stage:1cell	GSM5509227	GSM5509227: tRNA 1cell; Danio rerio; RNA Seq	GSM5509227		1	About 50 100 zebrafish eggs or embryos at a desired stage were collected to extract total RNA and dechorionated by Pronase digestion. Embryos were then transferred to 1.5 ml eppendorf tube and lysed in 1 ml Trizol Reagent Thermo Fisher Scientific 15596018. The lysate was centrifuged at 12 000 rpm for 10 min and the supernatant was collected and mixed with 200 μl chloroform in a fresh tube. post centrifugation at 12 000 rpm for 10 min at 4℃ the supernatant was transferred to a fresh tube with addition of equal volume of isopropanol and incubated at room temperature for 15 min followed by thorough mix and centrifugation at 12 000 rpm for 10 20 min. The RNA pellet was washed by 75% ethanol once and was stored as pellet in 75% ethanol at 80℃ until all samples were collected. Total RNA was placed on ice for 30 min with 10% PEG8000 and 0.5 M NaCl then centrifuged at 14 000 rpm for 15 min at 4℃. Supernatant low molecular weight LMW RNA was collected and precipitated by addition of ethanol. The LMW RNA was separated by 8% 8 M Urea PAGE gel then the bands correspond to 60 120 nt length were cut and crunched into small pieces. RNA was eluted from the gel by 0.3 M NaOAc overnight on a vertical rotator at 4℃. Then the supernatant was precipitated at 20℃ post addition with 2 μl of Glycoblue and 2.5 volume of ethanol. post centrifuged at 12 000 rpm for 30 min and washed by 75% ethanol the RNA precipitant was dissolved in RNase free water. There are three pretreatment steps before library preparation. 60 120nt RNA enriched in the previous step was first incubated in 100 mM Tris HCl pH 9.0 at 37℃ for 30 min to be deacylated then the RNA was treated with T4 PNK to repair ends at 37℃ for 30 min. Modifications on tRNAs were removed by ALKB treatment [1 μg purified RNA was treated by ALKB protein mixture wt ALKB 160 pmol; ALKB D135S 200 pmol; and ALKB D135S/L118V 200 pmol in reaction buffer 300 mM KCl 2 mM MgCl2 50 μM NH42FeSO42 300 μM α ketoglutarate 2 mM vitamin C 50 μg/ml BSA 50 mM MES pH 5.0 40 U RNase Inhibitor 1x Protease Inhibitor for 2 h at room temperature]. Purification was needed post each pre treatment step. 100 ng RNA was then subjected for RNA library preparation. tRNA library was prepared by VAHTS Small RNA Library Prep Kit for Illumina Vazyme NR801. RNA was heat denatured at 80℃ for 2 min in the first step then every steps were done followed the manual in kit. The libraries were first concentrated by DNA concentrator spin column Zymo then separated on 6% native PAGE gel. Bands correspond to 180 240 bp 60 120 nt RNA ligated with both adaptors length were cut and DNA libraries were eluted from gels.	GEO Accession:GSM5509227	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000		SRP331635			tRNA_1cell_1.fq.gz tRNA_1cell_2.fq.gz	fastq fastq	9361966500.0	31206555.0	GSM5509227 r1	0:150 1:150	A:1533392527;C:1543333302;G:4733567814;T:1550956628;N:716229	150	150			1533392527	1543333302	4733567814	1550956628	716229	SRX11676000	SRS9707982	SRA1274130	GEO	Anming Meng Lab, School of Life Science, Tsinghua University	2	0.7668	0.00334	0.06232	0.00092	0.81858	0.99809	0.57209	0.77622	150	150	B	T	mate2 technical by mapping diff	illumina	novaseq_era	unknown	small_rna	unknown	bulk	unknown	unknown		China	2021-08-09	Zygote	Embryo	Whole Organism	All anatomical structures