|
| Status |
Public on Jul 02, 2021 |
| Title |
Testis - wt - smRNA-seq_rep3 |
| Sample type |
SRA |
| |
|
| Source name |
Testis - wt - smRNA-seq
|
| Organism |
Mesocricetus auratus |
| Characteristics |
tissue: Testis
genotype: w/w
|
| Treatment protocol |
PIWIL1 and PIWIL3 mutant golden hamster was generated and established using CRISPR/Cas9 system. See reference for details (Hasuwa et al.).
|
| Growth protocol |
Golden hamsters were purchased from Japan SLC and maintained with 14 light and 10 dark cycle. All animal experiments were approved by the Animal Care and Use Committee of the Keio university.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Mature females were induced to superovulate by i.p. injection of 40 IU PMSG (ASKA Animal Health) at 1200 to 1400 on the day of conspicuous, postestrus vaginal discharge (day 1 of the estrous cycle). GV oocytes were collected from ovary after 102h PMSG injection. MII oocytes were collected from oviduct after 115h PMSG injection. PN zygotes were collected from oviduct after 91 or 115h PMSG injection followed by mating with male during the night of day4 or 5 (depend on the female estrus cycle).
small-RNA-seq: Total RNAs for small RNA sequence were extracted from MII oocytes of PIWIL1 and PIWIL3 homozygous and heterozygous mutants or testis of PIWIL1 homozygous mutant and wild type. The construction of small RNA libraries was performed using NEXTFLEX® Small RNA-Seq Kit v3 (PerkinElmer) with the manufacturer’s instructions.
|
| |
|
| Library strategy |
ncRNA-Seq |
| Library source |
transcriptomic |
| Library selection |
size fractionation |
| Instrument model |
Illumina MiSeq |
| |
|
| Description |
TestisPIWIL1piRNA_count.tsv
|
| Data processing |
small-RNA-seq: The adapter sequence and 4nt random sequences at 3’ and 5’ ends were removed cutadapt v1.14. A quality filter of 30 was adopted by removing sequences below this threshold. Reads at the range of 10 ~ 40nt after adapter-removal were used for further analysis. Reads were mapped to the hamster reference genome (hamster.sequel.draft-20200302.arrow.fasta; 3) using Bowtie (v1.1.1), permitting 0 mismatches, and genome-mapped reads were used for further analysis. Genome mapped reads were compared between replicate samples and combined due to their high reproducibility. Reads were further mapped to tRNA, rRNA, sn/snoRNA, and miRNA. The read count mapped to miRNAs were used to normalize small RNA reads obtained from heterozygous and homozygous mutants. Read counts mapped to each unique small RNA sequence were compared between heterozygous and homozygous samples, and those with an 4-fold decrease and identical to previously described PIWI-associated piRNAs were extracted as PIWIL1- or PIWIL3-piRNA.
Genome_build: hamster.sequel.draft-20200302.arrow.fasta (newly sequenced genome available under accession number PRJDB10770 in DDBJ)
Supplementary_files_format_and_content: small-RNA-seq: each unique PIWIL1/PIWIL3-piRNA sequence (see reference for the defenition of PIWIL1/PIWIL3-piRNA) with count of obtained reads in tab separated format.
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| |
|
| Submission date |
Apr 22, 2021 |
| Last update date |
Jul 03, 2021 |
| Contact name |
Yuka W. Iwasaki |
| E-mail(s) |
iwasaki@keio.jp
|
| Organization name |
Keio University School of Medicine
|
| Department |
Department of Molecular Biology
|
| Lab |
Siomi Lab
|
| Street address |
35 Shinanomachi, Shinjuku-ku,
|
| City |
Tokyo |
| ZIP/Postal code |
160-8582 |
| Country |
Japan |
| |
|
| Platform ID |
GPL29575 |
| Series (1) |
| GSE164356 |
Production of functional oocytes requires maternally expressed PIWI genes and piRNAs in golden hamsters |
|
| Relations |
| BioSample |
SAMN18827153 |
| SRA |
SRX10655155 |
