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Sample GSM6372284

Query DataSets for GSM6372284

Status

Public on Jun 20, 2024

Title

siCTRL_rep2_24h

Sample type

SRA

Source name

HEK293T cell line

Organism

Homo sapiens

Characteristics

cell line: HEK293T
sirna: siCTRL
time point: 24h

Extracted molecule

polyA RNA

Extraction protocol

RNA was extracted from the HEK293T and C4-2 cells.

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

BGISEQ-500

Data processing

mRNA half-life of the HEK293T cells and C4-2 cells was computed as described before. In short, the mRNA levels were first normalized to RPKM (Reads Per Kilobase Million). To reduce the noise due to genes that showed a higher expression levels after inhibition of transcription, we then scaled the FPKM values based on the expression levels of top 10 genes which were most stable between time points, so that these genes will have no change of expression between time points. The RNA degradation rate kdecay was defined as the mean values of ratio from RPKM at time 0 over the RPKM value at each other time point (0hour, 6hour, 12hour, 24hour). After that the pseudo-mRNA half-life t1/2 was calculated as ln2/kdecay.
After base-calling using guppy_basecaller (version 3.6.1), the direct RNA-seq of Nanopore reads with quality value more than 7 were aligned to the human genome (Hg38, GRCh38.p13 from GENCODE database) using Minimap with --secondary=no -ax splice -uf -k14 option. Tombo resquiggle was used to assigned to transcript sequence as reference from the website: https://www.gencodegenes.org/human . We extracted the median, standard deviation, mean, and dwell time from Nanopore raw signal data in each direct RNA sequencing read to calculate the probability of Nm modification using the training models from rRNA. The modified base supported by at least 5 modified transcripts were identified as modified Nm sites.
Assembly: hg19 for NGS RNA-seq. hg38 for Nanopore RNA-seq.
Supplementary files format and content: *.count.txt: Tab-delimited text files include abundance measurements.
Supplementary files format and content: *.bw: bigWig files.

Submission date

Jul 21, 2022

Last update date

Jun 20, 2024

Contact name

Yanqiang Li

Organization name

Boston Children's Hospital

Department

Cardiology Department

Lab

Kaifu Chen Lab