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Sample GSM1303470 Query DataSets for GSM1303470
Status Public on Dec 17, 2014
Title hom RNF17 6 week old adult testis rep2 RNAseq
Sample type SRA
Source name adult testes of 6 week old mouse
Organism Mus musculus
Characteristics age: 6 weeks old
genotype/variation: homozygous RNF17
strain: C57BL/6
antibody: none
Treatment protocol IP with RNF17 antibodies were done according (Aravin et al. 2008 Mol. Cell). Manually dissected testes were homogenized in lyses buffer and diluted 5 times in IP buffer NT2 with RNF17 antibodies diluted 1:500 and incubated for 10 hours at 4C. Then protein A agarose beads (Roche) were added and incubated for 2 more hours at 4C. After 5 washes with NT2 buffer total RNA from immunoprecepitates was isolated by proteinase K treatment with following Phenol/Chloroform pH 4.8 (Ambion) extraction.
Growth protocol mice were maintained according to the guidelines of the Cold Spring Harbor Laboratory Institutional Animal Care and Use Committee.
Extracted molecule total RNA
Extraction protocol total RNA was prepared with Trizol reagent (Invitrogen)
Transcriptome libraries were prepared according (Armour et al. 2009, Nat Methods) using not-so-random priming (NSR). 1 ug total RNA was revere transcribed using SuperScript III enzyme with first-strand NSR primer. RNA template was removed with RNase H (Invitrogen) Then cDNA was mixed with exo− Klenow fragment (NEB) second-strand NSR primer to synthesize the second strand. For PCR amplification, purified second-strand synthesis reaction were mixed with ExpandPLUS enzyme (Roche) and PE-P5-SBS3 and PE-P7-SBS8 primers. Products were run on a 2% low-melt agarose gel, and the 350–500 bp. range were purified.
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina MiSeq
Data processing First 8 nucleotides from 5 'end were trimmed and reverse-complemented before genomic mapping
Mapped to mm9 mouse genome release with STAR (Dobin et al.29;15-21 2013 Bioinformatics) with 2 mismatches and maximum of 100 multiple alignments.
DESeq was used according to (Anders and Huber 2010, Genome Biol 11: R106.)
Genome_build: mm9
Supplementary_files_format_and_content: results of DESeq analysis only for transcripts which are deferentially expressed between homozygous and heterozygous RNF17mutant 6 week old adult testes include fold change in raw counts normalized according to DESeq protocol (Anders and Huber 2010, Genome Biol 11:R106) (up- (fold change<0) or down-regulated (fold change>0)) and p-values. Number of reads that map to genes or uniquely or multiple times (2-99) to tranposable elements form RNF17 IP or total testes lysate.
Submission date Jan 08, 2014
Last update date May 15, 2019
Contact name Vasily Vagin
Organization name Cold Spring Harbor
Lab Hannon's
Street address 1 Bungtown Rd
City Cold Spring Harbor
State/province NY
ZIP/Postal code 11724
Country USA
Platform ID GPL16417
Series (2)
GSE53913 RNF17 referees ping-pong in mouse testes [RNAseq]
GSE53919 RNF17 blocks promiscuous activity of PIWI proteins in mouse testes
BioSample SAMN02571037
SRA SRX423929

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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