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Sample GSM4471634 Query DataSets for GSM4471634
Status Public on Jul 23, 2020
Sample type SRA
Source name HCT-116
Organism Homo sapiens
Characteristics cell line: HCT-116
tissue: colorectal cancer
Treatment protocol Approximately 500,000 HCT-116 cells were plated in a single well of a 6-well plate. Cells were transfected with 2.5 µg of the SP1-PBase plasmid and 2.5 µg of the PB-SRT-Puro plasmid using Lipofectamine 3000 following manufacturer’s instructions. After 24 hours, cells were split and plated 1:10 in each of three 10 cm dishes. Puromycin was then added to a final concentration of 2 µg/ml and colonies were grown under selection for two weeks. We obtained approximately 2,300 colonies. All cells were pooled together and split into two populations. One half was subjected to DNA extraction, self-ligation, and inverse PCR, as described previously (Wang et al., 2012a), with the following modification: digestion with MspI was not performed as the SRT construct contained an second MspI cut site near the terminal repeat. The other half of cells underwent RNA extraction and SRT library preparation.
Growth protocol Cells were cultured in Dulbecco’s Modified Eagle Medium (DMEM) supplemented with 10% fetal bovine serum (FBS) and 1% of either penicillin-streptomycin or antibiotic-antimycotic. Cells were grown at 37ºC with 5% carbon dioxide (CO2). Media was replenished every 2 days.
Extracted molecule genomic DNA
Extraction protocol DNA extraction was performed as described previously (Wang et al., 2012a)
Self-ligation of digested genomic DNA and inverse PCR was performed as described previously (Wang et al., 2012a), with the following modification: digestion with MspI was not performed as the SRT construct contained an second MspI cut site near the terminal repeat.
Library strategy OTHER
Library source genomic
Library selection other
Instrument model Illumina NextSeq 500
Description Bulk DNA calling cards library
Data processing Reads were demultiplexed by the 3 base-pair barcode TAG followed by the end of the transposon terminal repeat, culminating with the piggyBac insertion site motif TTAA. Reads that had exact matches to these sequences were hard trimmed using cutadapt (Martin, 2011) with the following settings: -g "^TAGTTTACGCAGACTATCTTTCTAGGGTTAA" --minimum-length 1 --discard-untrimmed -e 0 --no-indels. Reads passing this filter were then trimmed of vector sequence along read 2 using cutadapt with the following settings: -g "^ATCACTTAAGCCGGTAC"" --minimum-length 1 --discard-untrimmed -e 0 --no-indels. The remaining reads were aligned to the human genome (build hg38) with NovoAlign and the following settings: -n 40 -o SAM -o SoftClip. Aligned reads were validated by confirming that they mapped adjacent to the insertion site motif. Successful reads were then converted to calling card format (.ccf.txt; see using custom programs (available at and visualized on the WashU Epigenome Browser v46 (Zhou et al., 2011) (
Genome_build: hg38
Supplementary_files_format_and_content: Supplementary_files_format_and_content: BAM files contain aligned and annotated calling card reads. ccf.txt files contain processed calling card data. describes calling card-specific BAM tags.
Submission date Apr 10, 2020
Last update date Jul 23, 2020
Contact name Robi D Mitra
Organization name Washington University in St. Louis
Department Genetics
Street address 4515 McKinley Ave
City St. Louis
State/province Missouri
ZIP/Postal code 63143
Country USA
Platform ID GPL18573
Series (1)
GSE148448 Self-reporting transposons enable simultaneous readout of gene expression and transcription factor binding in single cells
BioSample SAMN14573307
SRA SRX8094399

Supplementary file Size Download File type/resource
GSM4471634_DNA_v_RNA_DNACC.bam 397.0 Mb (ftp)(http) BAM
GSM4471634_DNA_v_RNA_DNACC.ccf.txt.gz 265.9 Kb (ftp)(http) TXT
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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