|
| Status |
Public on Oct 15, 2021 |
| Title |
RNAseq - LS180 cells, GFP_dox_neg_rep2 |
| Sample type |
SRA |
| |
|
| Source name |
LS180
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: Colorectal Cancer cells
cell line: LS180
doxycycline induction: -
expression: No doxycycline induced overexpression of GFP control
|
| Treatment protocol |
GFP, SOX9 and mutant SOX9 was induced by doxycycline (0.5 µg/ml) for 24h. Each experiment was performed in duplicates.
|
| Growth protocol |
LS180 cell line was obtained from Millipore Sigma and cultured in DMEM + 2mM Glutamine + 15% Fetal Bovine Serum (FBS) + 1% (P/S).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using RNeasy Mini kit (Qiagen) according to the manufacture`s instruction.
Matured mRNA fraction was captured and enriched via oligo(dT) beads, then mRNAs were fragmented randomly in fragmentation buffer and the first-strand cDNA is synthesized using random hexamer primers and M-MuLV Reverse Transcriptase (RNase H-). The second strand was subsequently generated by dNTPs, DNA polymerase I and RNase H. Double-stranded cDNA molecules were purified by AMPure XP beads and overhanging ends are repaired to blunt ends by exonuclease/polymerase. After 5’ phosphorylation and 3’ adenylation, the cDNAs were ligated with P5/P7 sequencing adapters, then size selection (insert fragment of preferentially 150~200 bp in length) and library purification was performed using AMPure XP system (Beckman Coulter, Beverly, USA). The final library was created by amplification and subsequent AMPure XP beads purification.
RNA-Seq was performed using 150-bp paired-end sequencing chemistry (Illumina) at Novogene core facility.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Data processing |
Illumina Casava1.7 software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to hg38 whole genome using STAR 2.7.0a
A gene count matrix was obtained using featureCounts on bam files
Genome_build: hg38
Supplementary_files_format_and_content: featureCounts_HT115.txt
Supplementary_files_format_and_content: normalized_counts.xls
|
| |
|
| Submission date |
Oct 12, 2021 |
| Last update date |
Oct 15, 2021 |
| Contact name |
Nilay S Sethi |
| E-mail(s) |
Nilay_sethi@dfci.harvard.edu
|
| Organization name |
Dana Farber Cancer Institute
|
| Department |
Department of Medical Oncology
|
| Lab |
Gastrointestinal Cancer Research
|
| Street address |
450 Brookline Ave
|
| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02215 |
| Country |
USA |
| |
|
| Platform ID |
GPL24676 |
| Series (2) |
| GSE185745 |
An enhancer-driven stem cell-like program mediated by SOX9 blocks intestinal differentiation in colorectal cancer [rnaseq_ls180] |
| GSE185747 |
An enhancer-driven stem cell-like program mediated by SOX9 blocks intestinal differentiation in colorectal cancer |
|
| Relations |
| BioSample |
SAMN22228177 |
| SRA |
SRX12579813 |
