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| Status |
Public on Jun 03, 2024 |
| Title |
WT-rep2 [RiboMeth-seq] |
| Sample type |
SRA |
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| Source name |
bone marrow cells
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| Organism |
Mus musculus |
| Characteristics |
tissue: bone marrow
genotype: wildtype
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| Extracted molecule |
total RNA |
| Extraction protocol |
Briefly, 1μg total RNA was subjected to alkaline hydrolysis, which was performed in 50 mM bicarbonate buffer pH 9.2 for 12 min at 95℃. The reaction was stopped by ethanol precipitation using 3M NaOAc (Sigma), pH 5.2 and Glycoblue (Invitrogen) as a carrier in liquid nitrogen. The size of generated RNA fragments was assessed by electrophoresis (1% agarose gel, 80V, 50 min) and were ranging from 50 to 200 nt. RNA fragments were directly 3’-end dephosphorylated using 5 U Antarctic Phosphatase (NEB) for 30 min at 37℃, and then phosphorylated at the 5’-end using T4 PNK and 1mM ATP for 1h at 37℃.Resulting RNA fragments were then purified using Rneasy MinElute Cleanup kit (QIAGEN) according to the manufacturer instructions with minor modification. Finally, 10 μL of nuclease-free water was used to elute RNA fragments.
Construction of RNA fragments library was performed by LC Sciences (China) using TruSeq Small RNA Sample Prep Kits (Illumina) following manufacturer instruction. After PCR amplification, products were purified on a 6% poly-acrylamide Tris-borate-EDTA gel. DNA fragments corresponding to 140~270 bp (the length of fragments plus adaptors) were recovered, and were sequenced on Illumina Hiseq2500 platform and single-end 50 bp reads were generated.
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| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 2500 |
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| Data processing |
Library strategy: RiboMeth-seq
Illumina Casava1.8 software used for basecalling.
Sequenced data were trimmed with trimmomatic and keep reads which length more than 16bp.
Clean data were mapped to rRNA reference (NR_003278.3 18S rRNA, NR_003280.2 5.8S rRNA, NR_003279.1 28S rRNA) using bowtie2 with parameter “--end-to-end -k 1”.
The sam files obtained from the alignment were converted to the bed files.
5’ and 3’ ends counting was done by bedtools (2.29.0) and combined to calculate RiboMeth score. The scores were normalized to average values for -6 and +6 nucleotides.
Genome_build: GRCm38
Supplementary_files_format_and_content: BedGraph files with each sample RiboMeth score.
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| Submission date |
Feb 10, 2021 |
| Last update date |
Jun 03, 2024 |
| Contact name |
Jiang Penglei |
| E-mail(s) |
jpl3982233@zju.edu.cn
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| Organization name |
Zhejiang University
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| Department |
School of basic medicine science
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| Street address |
Yuhangtang Road 866
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| City |
Hangzhou |
| ZIP/Postal code |
310058 |
| Country |
China |
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| Platform ID |
GPL17021 |
| Series (2) |
| GSE166515 |
Small Nucleolar RNAs in the Dlk1-Gtl2 Imprinting Locus Maintains LT-HSC Self-renewal by Regulation of rRNA Modification and Translation [RiboMeth-seq] |
| GSE166518 |
Small Nucleolar RNAs in the Dlk1-Gtl2 Imprinting Locus Maintains LT-HSC Self-renewal by Regulation of rRNA Modification and Translation |
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| Relations |
| BioSample |
SAMN17850041 |
| SRA |
SRX10064672 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM5073634_WT2.methscore.bedGraph.gz |
78.9 Kb |
(ftp)(http) |
BEDGRAPH |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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