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Sample GSM5011955 Query DataSets for GSM5011955
Status Public on Apr 14, 2021
Title Sample 120_Plate 2_Negative control (H2O)_Rep 2
Sample type SRA
Source name Plate 2_Negative control (H2O)
Organism blank sample
Characteristics sample type: Water; negative control
Treatment protocol Prior to exposure, cells were plated in 6-well plates with 5 mL of 3x10^5 cells/mL suspension in each well. Cells were exposed to each chemical prepared in their respective vehicle solvent (1% v/v). After 4 hours, cells were rinsed once with PBS and lysed in TempO-Seq lysis buffer. RNA was extracted using a Qiagen RNeasy Mini kit. The exposure was performed on three separate occasions to produce three replicates of each sample.
Growth protocol TK6 cells were cultured in suspension in a T75 flask in RPMI1640 medium supplemented with 10% v/v heat-inactivated horse serum, 2 mM L-glutamine, and 1 mM sodium pyruvate at 37ºC in 5% CO2. The cell density was maintained between 1x10^5 cells/mL and 1x10^6 cells/mL.
Extracted molecule total RNA
Extraction protocol Total RNA was extracted using a Qiagen RNeasy Mini kit. Quality was assessed using High Sensitiy RNA ScreenTapes on an Agilent TapeStation. RNA integrity number (RIN) of all samples was above 7.5.
The Templated Oligo-Sequencing (TempO-Seq) Human Whole Transcriptome Assay kit (BioSpyder, Carlsbad, CA) was used to generate sequencing libraries using total RNA from TK6 cells following the manufacturer's protocol. An input of 100ng was used from each RNA sample.
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina NextSeq 500
Description Plate #2 control
Data processing library strategy: TempO-Seq
Initial quality assessment was performed using negative controls (water only) and reference standards (human universal reference RNA and human brain total RNA) included on every TempO-Seq plate. We confirmed very little signal (< 0.3%) from samples with no cells/RNA. The BCL files were converted to FASTQ files and the reads were demultiplexed using bcl2fastq v. The FASTQ files were processed using the “pete. star. Script_v3.0” (BioSpyder). The script uses star v.2.5 and the qCount function from QuasR to align the reads and to extract the feature counts specified in a Gene Transfer Format (GTF) file from the aligned reads, respectively.
Using R 3.4.1, the read counts of all samples were normalized as counts per million (CPM) (Law et al. 2014).
Supplementary_files_format_and_content: Text file with counts per million (CPM) for each gene
Submission date Jan 08, 2021
Last update date Apr 14, 2021
Contact name Matthew Meier
Phone 613-447-6705
Organization name Government of Canada
Department Health Canada
Lab Mechanistic Studies Division Genomics Laboratory
Street address 251 Sir Frederick Banting Driveway
City Ottawa
State/province Ontario
ZIP/Postal code K1A 0K9
Country Canada
Platform ID GPL29107
Series (1)
GSE164478 Development of a transcriptomic biomarker to detect histone deacetylase inhibitors (HDACi) in TK6 cells
BioSample SAMN17265061
SRA SRX9808462

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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