|
| Status |
Public on May 10, 2019 |
| Title |
RNA-seq_HAP1_SMARCC2_r2 |
| Sample type |
SRA |
| |
|
| Source name |
HAP1 cell line
|
| Organism |
Homo sapiens |
| Characteristics |
library: RNA-seq
cell line: HAP1
replicate: 2
knockout: SMARCC2
|
| Growth protocol |
HAP1 WT and knock-out cells were cultured in Iscove’s Modified Dulbecco’s Medium (IMDM, Thermo Fisher Scientific, 21980-032) supplemented with 10% heat inactivated Fetal Bovine Serum (FBS, Thermo Fisher Scientific, 10500) and 1% Pen Strep (100 units/ml Penicillin, 100 µg/ml Streptomycin, Thermo Fisher Scientific, 15140-122). A549 cells were cultured in F-12K Nut Mix (Thermo Fisher Scientific, 21127-022) supplemented with 10% FBS and 1% Pen Strep.
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
RNA extraction was performed with Qiagen RNeasy Mini Kit (Cat No. 74106) according to the manufacturer's instructions.
Total RNA was quantified using Qubit 2.0 Fluorometric Quantitation system (Life Technologies) and the RNA integrity number (RIN) was determined using Experion Automated Electrophoresis System (Bio-Rad). RNA-seq libraries were prepared with TruSeq Stranded mRNA LT sample preparation kit (Illumina) using Sciclone and Zephyr liquid handling robotics (PerkinElmer). Library amount was quantified using Qubit 2.0 Fluorometric Quantitation system (Life Technologies) and the size distribution was assessed using Experion Automated Electrophoresis System (Bio-Rad). For sequencing libraries were pooled, diluted and sequenced on Illumina HiSeq 3000 using 50 bp single-read chemistry.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 4000 |
| |
|
| Description |
RNA-seq on HAP1 cell line with SMARCC2 KO, replicate 2
baf_complex.rnaseq.expression_counts.gene_level.csv
|
| Data processing |
Base calls provided by the Illumina Realtime Analysis software were converted into BAM format using Illumina2bam and demultiplexed using BamIndexDecoder (https://github.com/wtsi-npg/illumina2bam)
Reads were trimmed with Trimmomatic and aligned to the GRCh37/hg19 assembly of the human genome using Bowtie1 with the following parameters: "-q -p 6 -a -m 100-minins 0-maxins 5000-fr-sam-chunkmbs 200".
Duplicate reads were removed with Picard’s MarkDuplicates utility with standard parameters before transcript quantification with BitSeq using the Markov chain Monte Carlo method and standard parameters.
To obtain gene-level quantifications, we assigned the expression values of its highest expressed transcript to each gene.
Differential gene-level expression between each knockout and wild-type cells was performed with DESeq2 from the raw count data with a significance threshold of 0.05.
Genome_build: hg19
Supplementary_files_format_and_content: Each csv file contains read counts for each sample at gene level
|
| |
|
| Submission date |
May 09, 2019 |
| Last update date |
May 10, 2019 |
| Contact name |
Christoph Bock |
| E-mail(s) |
cbock@cemm.oeaw.ac.at
|
| Organization name |
CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences
|
| Street address |
Lazarettgasse 14
|
| City |
Vienna |
| ZIP/Postal code |
1090 |
| Country |
Austria |
| |
|
| Platform ID |
GPL20301 |
| Series (1) |
| GSE108388 |
Systematic functional characterization of BAF mutations yields novel intra-complex synthetic lethalities [RNA-Seq] |
|
| Relations |
| BioSample |
SAMN11611954 |
| SRA |
SRX5812542 |
