|
| Status |
Public on Aug 01, 2021 |
| Title |
HeLa_small_RNA_shAlkBH1_treated.rep1 |
| Sample type |
SRA |
| |
|
| Source name |
HeLa cells
|
| Organism |
Homo sapiens |
| Characteristics |
genotype: AlkBH1 knockdown
cell line: HeLa
rna population: small RNA
|
| Growth protocol |
HeLa cells were cultured in DMEM (Gibco 11995) supplemented with 10% (v/v) fetal bovine serum (Gibco), 1% penicillin and streptomycin (Gibco) and grown at 37 ˚C with 5% CO2
|
| Extracted molecule |
total RNA |
| Extraction protocol |
200-300 ng of cellular small RNA (size < 200 nt) was fragmented into 40-50 nt using RNA Fragmentation Reagent (AM8740, Invitrogen) following the manufacturer’s protocol and purified by Oligo Clean & Concentrator. Then RNA was subjected to demethylation treatment with 1.0 µL of engineered AlkB (D135S, 10 mg/mL) with the reported recipe, followed by purification using Oligo Clean & Concentrator.
Libraries were prepared according to Illumina's instructions
|
| |
|
| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Data processing |
Basecalls performed using CASAVA version 1.4
Low-quality and adapter-containing reads were trimmed from Spike-In oligos raw sequencing data using trim-galore package in single-end mode. Reads shorter than 50 bp were removed.
We used fastx_collapser in FASTX-toolkit to collapse identical sequences in a fastq file into a single sequence, which was used to remove duplicated reads from PCR amplification.
The modified reads were aligned to the reference genome (hg19 for HeLa cells or mm10 for mESC) using bowtie2 (v2.3.3.1)(small RNA) and tophat2(PolyA+ RNA) under default parameters.
Mapped sam files were subsequently converted and sorted to bam files using samtools sort (v1.9). Sorted bam files were subsequently filtered to get the unique mapped reads using samtools view(-q 5). Rnaseqmut was used to identify mutations under the “-t -s 2 -m 2” parameters.
Genome_build: hg19
Supplementary_files_format_and_content: txt/mutation list
Library strategy: f5C-seq
|
| |
|
| Submission date |
Aug 26, 2020 |
| Last update date |
Aug 01, 2021 |
| Contact name |
Ruitu Lyu |
| E-mail(s) |
lvruitu@uchicago.edu
|
| Organization name |
The University of Chicago
|
| Department |
Chemistry
|
| Lab |
Chuan He
|
| Street address |
929 E 57th Street
|
| City |
Chicago |
| State/province |
IL |
| ZIP/Postal code |
60637 |
| Country |
USA |
| |
|
| Platform ID |
GPL24676 |
| Series (1) |
| GSE156933 |
A Quantitative Sequencing Method for 5-Formylcytosine in RNA |
|
| Relations |
| BioSample |
SAMN15914514 |
| SRA |
SRX9014813 |
