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| Status |
Public on Dec 31, 2014 |
| Title |
Flag-Siwi bound piRNAs from Vasa siRNA treated BmN4 |
| Sample type |
SRA |
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| Source name |
BmN4 cells
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| Organism |
Bombyx mori |
| Characteristics |
cell line: BmN4
treatment: Vasa siRNA
antibody: anti-Flag M2 (Sigma)
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| Treatment protocol |
BmN4 cells were collected and Immuno-precipitation was performed using Flag (for Flag-Siwi expressed in RNAi samples) antibody.
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| Growth protocol |
BmN4 cells were grown at 27 degrees on Ex-Cell medium
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| Extracted molecule |
total RNA |
| Extraction protocol |
Siwi, BmAgo3, and Flag-Siwi complexes were immunopurified from BmN4 cells (untreated or treated with indicated siRNAs) using anti-Siwi, anti-BmAgo3, and anti-Flag antibody bound to Dynabeads protein G (Invitrogen). The beads were washed three times with binding buffer [30 mM HEPES (pH 7.4), 150 mM potassium acetate, 5 mM magnesium acetate, 5 mM DTT, 0.1 % NP-40, 2 mg/mL pepstatin, 2 mg/mL leupeptin and 0.5% aprotinin] containing 500mM sodium chloride and twice with binding buffer. Total RNAs were eluted from the beads using phenol–chloroform and precipitated using ethanol.
small RNA-seq libraries were prepared using NEBNext Small RNA Library Sample Prep Set (New England BioLabs (NEB), E7330).
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| Library strategy |
RIP-Seq |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina MiSeq |
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| Description |
RNA (immunoprecipitated)
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| Data processing |
Sequencing of the libraries generated from Siwi, BmAgo3, and Flag-Siwi IP products were performed using MiSeq (illumina). Adapter sequences were removed from obtained reads, and mapped to the B. mori genomic sequence (Mita et al., 2004) using Bowtie2 (Langmead et al., 2012), allowing zero mismatches. The genome-mapped reads were aligned to B. mori transposable element consensus sequences (kind gift from Dr. Kawaoka) using Bowtie2 for further analysis.
Genome_build: genomic sequence from kaikobase (http://sgp.dna.affrc.go.jp/), integretedseq.txt.gz
Supplementary_files_format_and_content: tab-delimited text files include genome-mapped small RNA sequences and their read counts for small RNAs with 10 or more read counts
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| Submission date |
Oct 29, 2014 |
| Last update date |
Jul 03, 2021 |
| Contact name |
Yuka W. Iwasaki |
| E-mail(s) |
iwasaki@keio.jp
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| Organization name |
Keio University School of Medicine
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| Department |
Department of Molecular Biology
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| Lab |
Siomi Lab
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| Street address |
35 Shinanomachi, Shinjuku-ku,
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| City |
Tokyo |
| ZIP/Postal code |
160-8582 |
| Country |
Japan |
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| Platform ID |
GPL18754 |
| Series (1) |
| GSE58221 |
High throughput sequencing of Siwi and BmAGO3 associated piRNAs from Bombyx mori BmN4 cells |
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| Relations |
| BioSample |
SAMN03151497 |
| SRA |
SRX746147 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1533654_VasaRNAi_Siwi_piRNAs.txt.gz |
410.7 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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