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| Status |
Public on Nov 12, 2020 |
| Title |
12h-high-TNFa treatment RNAseq, rep2 |
| Sample type |
SRA |
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| Source name |
12h-high-TNFa treated HEK 293 cells
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| Organism |
Homo sapiens |
| Characteristics |
cell line: HEK 293 transfected with reporter construct
treatment: TNF-alpha treatment 12 hours, 50 ng/mL
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| Treatment protocol |
The final concentration of TNF (high, default) is 50 ng/mL.
The final concentration of TNF (low, specified) is 0.4 ng/mL.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA of cells was isolated using RNeasy Mini Kit (Qiagen), and cDNA were synthesized using PrimeScript first strand cDNA synthesis kit (Takara) with oligo-dT.
The RNA-seq libraries were constructed by BGI, Shenzhen, as the platform BGISEQ-500 instruction described.
ChIP-seq were performed as previously described (Xiong et al., Molecular Cell 64, 913-925).
The ChIP-seq libraries were constructed using NEBNext DNA Sample Prep Master Mix (NEB). DNA libraries were subjected to size-selection using AMPure XP beads (Beckman Coulter) and PCR amplification (18-20 cycles for ChIP-seq DNA libraries).
The NEBNext Enzymatic Methyl-seq Conversion Module (NEB E7125L) was used for measuring genome-wide DNA methylation level. Purified genomic DNA containing 0.05% CpG methylated pUC19 control DNA and 1% unmethylated lambda DNA was sheared to a mean length of 450 bp with Covaris M220 Focused-ultrasonicator. End repair, A-tailing, methylated adaptor ligation and size selection were performed on 125 ng fragmented genomic DNA with Kapa hyper prep kit (Kapa Biosystems KK8504) according to the manufacturer’s instructions. The methylated adaptor ligated DNA fragments were then treated with oxidation of 5mC and 5hmC, clean-up of TET2 converted DNA, denaturation of DNA, deamination of cytosines and clean-up of deaminated DNA using the NEBNext Enzymatic Methyl-seq Conversion Module (NEB E7125L). The deaminated DNA was amplified with KAPA HiFi Hotstart Uracil+ ReadyMix PCR kit using primers from NEBNext multiplex oligos for Illumina (index primers set 1) (NEB E7335) (Kapa Biosystems KK2801) according to the manufacturer’s protocol. PCR cycling condition was as follows: 98℃ for 45 s, followed by 7 cycles of 98℃ for 15 s, 60℃ for 30 s, 72℃ for 30 s, and final extension 72℃ for 1 min, then hold at 4℃. Libraries were sequenced on an Illumina NovaSeq 6000 using 150-bp paired-end mode.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
BGISEQ-500 |
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| Description |
embryonic kidney cells
somatic cell
MGISEQ-2000
mRNA
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| Data processing |
RNA-seq basecalls performed by BGI, Shenzhen.
RNA-Seq data were mapped to Human Gencode release 24 by STAR aligner and quantified by Cuffdiff.
ChIP-seq basecalls performed by Berry Genomics, Co. Ltd. Beijing.
ChIP reads were mapped to UCSC hg38 genome by Bowtie2 (2.3.4.2).
ChIP data were filtered using samtools rmdup to remove potential PCR duplicates.
Enzymatic Methyl-seq reads in PE150 were firstly assessed by FASTQC software and then trimmed by trim_galore to remove adapters and low-quality bases. The filtered reads were then mapped to human genome (hg38) using the Bismark software (v0.22.3). Duplicate reads were discarded, and methylation information were called by Bismark.
Genome_build: hg38 (GRCh38)
Supplementary_files_format_and_content: FPKM values were calculated by Cuffdiff and stored as tab-separated texts.
Supplementary_files_format_and_content: bigWig files were generated using IGV tools with fragment extension of average fragments size.
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| Submission date |
Aug 07, 2020 |
| Last update date |
Nov 14, 2020 |
| Contact name |
Zhuqiang Zhang |
| E-mail(s) |
[email protected]
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| Organization name |
Institute of Biophysics, CAS
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| Street address |
15 Datun Road, Chaoyang District.
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| City |
Beijing |
| State/province |
--- |
| ZIP/Postal code |
100101 |
| Country |
China |
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| Platform ID |
GPL23227 |
| Series (1) |
| GSE152146 |
TNF induced inflammatory transcription dynamics and epigenetic changes |
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| Relations |
| BioSample |
SAMN15760212 |
| SRA |
SRX8910124 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4715097_B50-rep2.txt.gz |
460.2 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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