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Status |
Public on Dec 20, 2019 |
Title |
RNAseq_B2B_Ni_washout_Dup |
Sample type |
SRA |
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Source name |
Nickel-washed-out cells (Ni-W)_replicate
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Organism |
Homo sapiens |
Characteristics |
cell line: BEAS-2B cell type: Human lung epithelial cell line treatment: NiCl2-100µM for 6 Weeks, following 6-week Ni exposure, single cells clone were isolated and expanded the populations to obtain homogenous clones of Ni-washed-out cells
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Treatment protocol |
Both BEAS-2B and RT4 cells cells were exposed to 100 μM NiCl2 for 6 weeks. Following exposure, the cells were washed and cultured in Ni‐free medium for 2 weeks to obtain Ni‐washed‐out cells. For BEAS-2B cells, following 6-week exposure, a homogenous populations of Ni-exposed cells was isolated as described. Briefly, the Ni-exposed cells were then plated in 15 cm plates at the rate of 1000, 500, 100 and 50 cells per plate in Ni-free medium. Nicely separated single colonies were randomly isolated and the populations were expanded in Ni-free medium. A randomly selected homogenous population of Ni-washed-out cells (Ni-W) in culture for >6 months post Ni exposure was used for a detailed examination.
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Growth protocol |
Human lung epithelial BEAS-2B cells (ATCC, CRL-9609) were cultured in Dulbecco’s Modified Eagle’s Medium (DMEM, Cellgro) supplemented with 1% Penicillin-Streptomycin and 10% Fetal Bovine Serum (FBS, Atlanta Biologicals) at 37°C and 5 % CO2. Human urinary bladder epithelial RT4 cells were cultured in McCoy's 5A Modified Medium (Life Technologies, Carlsbad, CA), supplemented with 10% FBS (Atlanta Biologicals) and 0.5% Penicillin‐Streptomycin at 37°C and 5% CO2.
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Extracted molecule |
total RNA |
Extraction protocol |
Total RNA was isolated using RNeasy Kit (Qiagen 74104). For ChIP the cells were crosslinked with 1% formaldehyde for 10 minutes at 25°C and sonicated to obtain 200-500bp fragments. ChIP was performed using ChIP grade antibodies. RNA-Seq libraries were prepared using Illumina TruSeq RNA Sample Preparation Kit (RS-122-2002) according to manufacturer’s protocol. ChIP-Seq libraries were prepared using Illumina TruSeq ChIP sample preparation Kit (IP -202-1024), according to manufacturer’s protocol.
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Library strategy |
RNA-Seq |
Library source |
transcriptomic |
Library selection |
cDNA |
Instrument model |
Illumina HiSeq 2500 |
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Description |
RNAseq_B2B_Ni_washout_Dup BEAS-2B cells treated with NiCl2-100µM for 6 Weeks, following 6-week Ni exposure, single cells clone were isolated and expanded the populations to obtain homogenous clones of Ni-washed-out cells.
|
Data processing |
Illumina Casava1.8 software used for basecalling.
RNA-Seq data analysis was performed using Biowardrobe experimental management system. Breifly Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to GRCh37/hg19 whole genome using STAR version 2.4.oj using default parameters; multi hits removed
Gene expression levels were quantified as reads per kilobase of exon per million fragments mapped (RPKM) using Biowardrobe algorithm
Differential gene expression was calculated using DESeq2
More information on Biowardrobe suite can be found at https://www.biowardrobe.com/
Genome_build: Feb. 2009 (GRCh37/hg19)
Supplementary_files_format_and_content: csv file include RPKM values for each Sample
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Submission date |
Aug 28, 2019 |
Last update date |
Dec 20, 2019 |
Contact name |
Rama Natarajan |
E-mail(s) |
RNatarajan@coh.org
|
Organization name |
Beckman Research Institute, City of Hope
|
Department |
Department of Diabetes Complications and Metabolism
|
Lab |
Rama Natarajan
|
Street address |
1500 E Duarte Rd
|
City |
Duarte |
State/province |
California |
ZIP/Postal code |
91010 |
Country |
USA |
|
|
Platform ID |
GPL16791 |
Series (2) |
GSE136558 |
Nickel induced transcriptional changes persist post exposure through epigenetic reprograming (RNA-seq dataset) |
GSE139356 |
Nickel induced transcriptional changes persist post exposure through epigenetic reprograming |
|
Relations |
BioSample |
SAMN12647558 |
SRA |
SRX6766551 |