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| Status |
Public on May 07, 2016 |
| Title |
human1_PS_RNA |
| Sample type |
SRA |
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| Source name |
adult testis
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| Organism |
Homo sapiens |
| Characteristics |
tissue: testis
cell type: pachytene spermatocytes
chip antibody: none
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| Growth protocol |
Human testis samples were obtained from patients undergoing vasectomy reversals at the Infertility Clinic of St. Louis.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Pachytene spermatocyte and round spermatid fractions were collected by StaPut (Bellve et al 1993). Purity was assessed under phase optics and was consistently >95% for each desired cell type. Cells were washed once in PBS and then split into two aliquots. One aliquot (for ChIP) was fixed in 1% formaldehyde then quenched with 2.5M glycine, while the second (for RNA) was left unfixed. Both fixed and unfixed aliquots were snap frozen in liquid nitrogen, then stored at -80C. Unfixed aliquots of sorted cells were thawed on ice, and RNA isolation was performed using the RNEasy Mini kit according to the manufacturer’s instructions. Genomic DNA was removed using gDNA eliminator columns supplied with the kit.
RNA-seq libraries were prepared using a SMARTer stranded RNA prep kit (Clontech) according to the manufacturer’s instructions.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
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| Data processing |
Image analysis and base calling was done using the standard Illumina pipeline for HiSeq2500.
Data was quality-filtered using fastq_quality_filter from the FASTX toolkit with the following parameters: -q 20 -p 80. For libraries with >50% adapter reads (one library total: human_PS_K4me3_ChIP), reads containing only adapter sequence were discarded using fastx_clipper.
For RNA-seq, FPKM values were generated using Cufflinks 2.2 with default parameters, except that a transcript gtf was supplied (-G option), and average fragment length of the library was supplied under the -m flag.
genome build: hg19
processed data files format and content: All files are tab-delimited text (.txt). For RNA-seq, FPKM is reported for each Ensembl (build 75) transcript and gene. For ChIP-seq, raw counts normalized to (aligned, non-duplicated) library size are reported for the +/- 2kb region surrounding each Ensembl (build 75) gene.
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| Submission date |
May 04, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Bluma J Lesch |
| Organization name |
Yale University
|
| Department |
Genetics
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| Street address |
333 Cedar Street
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| City |
New Haven |
| State/province |
CT |
| ZIP/Postal code |
06525 |
| Country |
USA |
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| Platform ID |
GPL16791 |
| Series (1) |
| GSE68507 |
Cross-species comparison of epigenetic poising in the mammalian germ line |
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| Relations |
| BioSample |
SAMN03481896 |
| SRA |
SRX994298 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1673959_human1_PS_FPKM.txt.gz |
1.9 Mb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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