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| Status |
Public on Dec 23, 2019 |
| Title |
Mouse-MIWI-HET-rep2 |
| Sample type |
SRA |
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| Source name |
Testis
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| Organism |
Mus musculus |
| Characteristics |
tissue: Testis
developmental stage: Adult
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from the mouse FACS-sorted germ cells using mirVana miRNA isolation kit (Thermo Fisher, AM1560).
In brief, total RNA was run on 15% urea polyacrylamide gel electrophoresis (PAGE) to select small RNAs ranging from 18–34 nt. To prevent microRNAs and siRNAs entering to the library, sRNAs were oxidized with sodium periodate (NaIO4) prior to 3′ adaptor ligation. We then ligated the 3′ adaptor (5′-rApp NNN TGG AAT TCT CGG GTG CCA AGG /ddC/-3′ or 5′-rApp TGG AAT TCT CGG GTG CCA AGG /ddC/-3′; Supplementary file 1) in the presence of 50% (w/v) PEG8000 (Rigaku, 1008063). After 3′ ligation, ligated sRNAs were run on 15% PAGE to eliminate the non-ligated 3′ adaptor; ligated sRNAs that were 42 to 60 nt with 3′ adaptor were purified. Thereafter, 5′ adaptor was ligated.
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| Library strategy |
ncRNA-Seq |
| Library source |
transcriptomic |
| Library selection |
size fractionation |
| Instrument model |
Illumina NextSeq 500 |
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| Data processing |
The 3′ adaptor sequence from the reads were removed and those reads whose PHRED score <5 was further filtered. The reads passed the quality check were mapped to human genome (hg19) and or mouse genome (mm10) using piPipes allowing 1 mismatch.
For RNA-seq, CAGE and PAS-seq, we first removed rRNA reads using bowtie 2.2.5 with default parameters48 and then the unaligned reads were further mapped to human genome (hg19) using STAR 2.3 (Ref. 52). Mapped results were generated as SAM format that was further transformed into duplication removed sorted BAM format using SAMtools 1.8 and normalized bigWig files are generated by custom scripts.
Raw reads were mapped to genome using Bowtie 2.2.5 with parameter —very-sensitive. Mapped results were generated as SAM format that was further transformed into duplication removed sorted BAM format using SAMtools 1.8. We, then, normalized genome coverage files in bigWig format that were calculated using homemade script. Data are presented in bigWig file of unique mapping reads as read depth per million reads (rpm). We used Model-based analysis of ChIP-seq (MACS 1.1.2; Ref. 58) with parameter -q 0.01 to detect A-MYB peaks that were significantly enriched over input (1,296 genomic regions; false discovery rate [FDR] < 0.05).
Genome_build: hg19; mm10
Supplementary_files_format_and_content: bigWig; narrowPeak; rpm.txt (text)
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| Submission date |
Aug 13, 2019 |
| Last update date |
Dec 23, 2019 |
| Contact name |
Tianxiong Yu |
| Organization name |
Umass Med
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| Street address |
364 Plantation St
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| City |
Worcester |
| State/province |
MA |
| ZIP/Postal code |
01605 |
| Country |
USA |
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| Platform ID |
GPL19057 |
| Series (1) |
| GSE135791 |
Pachytene piRNA Genes Are Rapidly Diverging Among Modern Humans |
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| Relations |
| BioSample |
SAMN10938300 |
| SRA |
SRX5376398 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4030265_SRS-MIWI-HET-4-May29-4N-740-10pri-unox-id8_S19.rpm.txt.gz |
3.6 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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