|
Status |
Public on Jan 04, 2016 |
Title |
rep2_siCDC73 |
Sample type |
SRA |
|
|
Source name |
IMEC cell line
|
Organism |
Homo sapiens |
Characteristics |
sirna: siCDC73 replicates: 2
|
Treatment protocol |
Cells were infected with the indicated doxycycline-inducible constructs and treated with Dox or EtOH as solvent control or transfected with the indicated siRNAs.
|
Growth protocol |
IMEC cell lines were grown in DMEM/F-12 with appropriate supplements.
|
Extracted molecule |
total RNA |
Extraction protocol |
Total RNA was extracted using the RNeasy Mini Kit (Qiagen) including on-column DNase digestion. Poly-A RNA was isolated from total RNA using the NEBNext Poly(A)mRNA Magnetic Isolation Module (E7490). For 4-SU seq, nascent RNA transcripts were labelled for 10 min using 1 mM 4-thiouridine (4sU, Sigma T4509). After RNA isolation the 4SU-labelled RNAs were biotinylated and enriched using MyOne Streptavidin beads (Life Technologies). Eluted RNAs were subsequently used for library preparation using NEBNext RNA-Sequencing kits. Libraries for the RNA-seq samples were constructed using the NEBNext Library Prep Kit (6100) following the instruction manual. Briefly, poly-A RNA was fragmented to generate 200 nucleotides long fragments. First and second strand cDNA synthesis was performed and the resulting cDNA was end-repaired, ligated to NEBNext adaptors. The cDNA was size selected using agarose gels and amplified by PCR. For the resulting RNA-seq libraries, amplicon sizes and quantities were determined using the Biorad Experion system. Libraries were sequenced on an Illumina NextSeq 500 following the manufacture’s instructions.
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|
|
Library strategy |
RNA-Seq |
Library source |
transcriptomic |
Library selection |
cDNA |
Instrument model |
Illumina NextSeq 500 |
|
|
Description |
polyA RNA norm_count_table_for_siMYC_siCDC73.csv
|
Data processing |
Basecalling and Demultiplexing was performed using BaseSpace from Illumina. Reads were aligned to the human genome (hg19) with BOWTIE2 v2.1.0 using standard-options Bedgraph files were generated using Bedtools Differential expression and statistical inference was done using edgeR. Genome_build: hg19 Supplementary_files_format_and_content: The csv-files contain a table showing the edgeR normalized counts for each transcript. The bedgraph files were generated using Bedtools for each sample.
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|
|
Submission date |
Nov 20, 2015 |
Last update date |
May 15, 2019 |
Contact name |
Martin Eilers |
Organization name |
University of Wuerzburg
|
Department |
Chair for Biochemistry and Molecular Biology
|
Lab |
Martin Eilers
|
Street address |
Am Hubland
|
City |
Wuerzburg |
ZIP/Postal code |
97074 |
Country |
Germany |
|
|
Platform ID |
GPL18573 |
Series (2) |
GSE70000 |
Ubiquitin-dependent turnover of MYC promotes loading of the PAF complex on RNA Polymerase II to drive transcriptional elongation (RNA-seq) |
GSE70009 |
Ubiquitin-dependent turnover of MYC promotes loading of the PAF complex on RNA Polymerase II to drive transcriptional elongation |
|
Relations |
BioSample |
SAMN04285082 |
SRA |
SRX1441822 |