NCBI Logo
GEO Logo
   NCBI > GEO > Accession DisplayHelp Not logged in | LoginHelp
GEO help: Mouse over screen elements for information.
          Go
Sample GSM6560796 Query DataSets for GSM6560796
Status Public on Sep 11, 2022
Title Cyclin B1 +ve [F09]
Sample type SRA
 
Source name colorectal tumour
Organism Homo sapiens
Characteristics tissue: colorectal tumour
cell line: COLO205
genotype: BRAF V600E
treatment: selumetinib (48h, 1uM)
Treatment protocol COLO205 cells were treated with 1uM selumetinib for 48 hours
Growth protocol COLO205 cell lines were provided by the laboratory of Dr. Simon J Cook (Babraham Institute). Cells were cultured in RPMI-1640 media supplemented with 10% (v/v) foetal bovine serum, penicillin (100 U/mL), streptomycin (100 mg/mL) and 2 mM glutamine at 37 °C in a humidified incubator with 5% (v/v) CO2.
Extracted molecule genomic DNA
Extraction protocol Cells were glyoxal-fixed and stained for CCNB1 as previously described (Channathodiyil and Houseley, 2021) and single cells sorted by flow cytometry to collect CCNB1 positive and CCNB1 negative cells in a 96-well plate.
Bisulfite conversion was carried out using EZ Methylation Direct bisulfite reagent (Zymo) and desulfonation and purification of converted DNA with magnetic beads (Zymo) on the Bravo automated workstation (Agilent) with DNA eluted into the master mix for the first-strand synthesis. Random priming and extension were performed followed by purification of the resulting DNA fragments. Second strand synthesis was then performed and the resulting fragments amplified with 14 PCR cycles. Amplified libraries were pooled (2 pools of 48 libraries each) and purified using 0.7× Ampure XP beads.
Single cell bisulfite libraries were then prepared as (CLark et al 2017)
 
Library strategy Bisulfite-Seq
Library source genomic
Library selection RANDOM
Instrument model Illumina NextSeq 500
 
Description Bisulfite converted
GEO report for rows E-H CCNB1 +ve.txt
Data processing adapter and quality trimming using Trim Galore (v0.6.2)
mapped to GRCh38 using Bismark v0.22.1
reads were quantified in 5 Mb windows spaced every 2.5 Mb across the genome, with windows of aberrantly high and low read count excluded
Log transformed datasets were normalised per probe by subtracting the median for that probe, which converts data to log2-fold change in copy number relative to average (the average copy number for each probe being the genomic copy number)
Enrichment normalisation for the 10th and 90th percentiles was then applied to each dataset normalise differences in total read count and distribution.
Assembly: GRCh38
Supplementary files format and content: Normalised total read counts for the CCNB1 negative and CCNB1 positive datasets are provided as separate files
 
Submission date Sep 08, 2022
Last update date Sep 12, 2022
Contact name Laura Biggins
E-mail(s) laura.biggins@babraham.ac.uk
Organization name The Babraham Institute
Department Bioinformatics
Street address Babraham Research Campus
City Cambridge
ZIP/Postal code CB22 3AT
Country United Kingdom
 
Platform ID GPL18573
Series (2)
GSE168604 Selumetinib
GSE212899 Epigenetic and genetic change through cell cycling under selumetinib treatment
Relations
BioSample SAMN30725294
SRA SRX17488121

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

| NLM | NIH | GEO Help | Disclaimer | Accessibility |
NCBI Home NCBI Search NCBI SiteMap