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Sample GSM4810499

Query DataSets for GSM4810499

Status

Public on Aug 08, 2021

Title

GA0hr_c

Sample type

SRA

Source name

HeLa_untreated

Organism

Homo sapiens

Characteristics

cell line: HeLa
cell type: human cervical cancer cells
treatment: untreated

Treatment protocol

Gallic acid was diluted into the cell culture medium to achieve the working concentration of 50 µg/mL, immediately before being applied to HeLa cells. The cells were then incubated with gallic acid for 0 hour (GA0hr, untreated), 2 hours (GA2hr), 4 hours (GA4hr), 6 hours (GA6hr), and 9 hours (GA9hr), before collected for RNA sequencing.

Growth protocol

HeLa cells were cultured in DMEM/F-12 (Dulbecco’s Modified Eagle medium: Nutrient Mixture F-12) media, supplemented with 10% fetal bovine serum (FBS), 2mM GlutaMAX, 100 U/mL penicillin, and 100 μg/mL streptomycin (Thermo Fisher Scientific, Carlsbad, CA, USA). The cells were maintained in optimal culture condition, at 37 degree Celsius (°C) under an atmosphere of 5% CO2. They were seeded on culture dishes for 24 hours to achieve 50–60% confluence, before subjected to experiment.

Extracted molecule

total RNA

Extraction protocol

The total RNA in these corresponding conditions was harvested using TRIzol Reagent (Thermo Fisher Scientific, Carlsbad, CA, USA). The total RNA was isolated and purified using the Direct-zol™ RNA MiniPrep (Qiagen, Irvine, CA, USA) with the standard protocol as described by the manufacturer.
The RNA was first subjected to the Ribosomal RNA depletion, which was performed with Ribo-zero Magnetic Gold Kit (Illumina Inc., San Diego, CA). Samples were randomly primed and fragmented based on manufacturer’s recommendation (NEBNext® Ultra™ II Directional RNA Library Prep Kit, Illumina). The first strand was synthesized with the Protoscript II Reverse Transcriptase with a longer extension period (30 minutes for 42°C). All remaining steps for library construction were performed according to the NEBNext® Ultra™ II Directional RNA Library Prep Kit for Illumina.

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina HiSeq 4000

Data processing

Illumina CASAVA v1.8 software was used for base calling
LncRNA-Seq reads were aligned to Homo sapiens (human) reference genome GRCh38 (hg38). Bowtie2 v2.2.8 software was used to build index of reference genome, and Paired-end clean reads of LncRNA-Seq were aligned to the reference genome using HISAT2 (Langmead, B.et al., Nat Methods. 2012 Mar 4;9(4):357-9) v2.0.4. HISAT2 was run with ‘--rna-strandness RF’. DEG analysis was done using Cuffdiff.
Raw data files were filtered with GATK standard filter method and other parameters ( cluster： 3；WindowSize: 35; QD < 2.0 or FS > 60.0 or MQ < 40.0 or SOR > 4.0 or MQRankSum < - 12.5 or ReadPosRankSum Ø -8.0 or DP < 10)
We have a quantitative analysis of mRNA and the screening lncRNA by cuffdiff software(http://cole-trapnell-lab.github.io/cufflinks/cuffdiff/index.html)to get the FPKM information of mRNA and lncRNA of every sample.
Genome_build: GRCh38 (hg38)
Supplementary_files_format_and_content: Text files include FPKM values for each of 15 samples, of all transcripts, mRNA transcripts, lncRNA transcripts and TUCP transcripts (as indicated in the processed data file names).

Submission date

Sep 30, 2020

Last update date

Aug 08, 2021

Contact name

Ho Man Holly Tang

E-mail(s)

hollytang@gmail.com

Organization name

The Chinese University of Hong Kong

Department

School of Life Sciences