|
| Status |
Public on Jun 07, 2018 |
| Title |
wt_pSpt5_rep2 |
| Sample type |
SRA |
| |
|
| Source name |
spt5-13myc::kanMX6 leu1-32 ura4-D18 his3-D1 ade6-M210 h+
|
| Organism |
Schizosaccharomyces pombe |
| Characteristics |
temperature c: 30
antibody: Anti-pSpt5 antibody (custom made from 21st Century Biochemicals, Marlboro, MA; Sanso et al. 2012)
assay: ChIP-seq
|
| Growth protocol |
ChIP-seq: S. pombe cultures were grown in YES media till OD600 ~ 0.4.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
ChIP-seq: After HCHO-crosslinking (15 min at 25°C) protein extract was prepared using Mini-beadbeater (Biospec; 30 sec "ON" and 30 sec "OFF"). Lysates were precleared with Protein A agarose beads and used for IP with respective antibodies.
ChIP-seq: Multiplexed ChIP-seq libraries were prepared using the Illumina TruSeq DNA Sample Preparation kit v2 with 75 ng of input or IP DNA and barcode adaptors.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Data processing |
ChIP-seq: Paired-end sequencing (50-nt reads) was performed on an Illumina HiSeq 2000 (Genome Quebec Innovation Centre, McGill University).
ChIP-seq: Quality control and adapter trimming of the FASTQ (raw sequencing files) files were done in Galaxy using FastQC Read Quality reports (Galaxy Version 0.69) and trimmomatic flexible read trimming tool for Illumina NGS data (Galaxy Version 0.36.3) respectively.
ChIP-seq: After adaptor trimming and quality control, processed FASTQ files were aligned to the S. pombe genome using Bowtie2 (Galaxy Version 2.2.6.2).
ChIP-seq: Aligned sequences of each biological replicate were fed into MACS2 (Galaxy Version 2.1.1.20160309.0) to call peaks from alignment results. Generated “bedgraph treatment” files were converted into bigwig using “Wig/BedGraph-to-bigWig converter” (Galaxy Version 1.1.0), concatenated (Galaxy Version 1.0.1) to combine replicates of each sample, subtracted IgG reads from each and processed using computeMatrix (Galaxy Version 2.3.6.0) in DeepTools to prepare data for plotting a heatmap and/ or a profile of given regions.
Genome_build: ChIP-seq: S. pombe: ASM294v2.
Supplementary_files_format_and_content: ChIP-seq: processed files are in bigWig format. There are 26-bigWig files in total. 4 for pSpt5, 4 for Myc, 3 for IgG and 2 for Iuput for each strain (wt or dis2cs) including two biological replicates.
|
| |
|
| Submission date |
Jan 12, 2018 |
| Last update date |
Jun 07, 2018 |
| Contact name |
Robert P Fisher |
| E-mail(s) |
robert.fisher@mssm.edu
|
| Phone |
2126598677
|
| Organization name |
Icahn School of Medicine at Mount Sinai
|
| Department |
Oncological Sciences
|
| Lab |
15-76
|
| Street address |
One Gustave L. Levy Place, Box 1130, L15-76
|
| City |
New York |
| State/province |
New York |
| ZIP/Postal code |
10029 |
| Country |
USA |
| |
|
| Platform ID |
GPL20584 |
| Series (1) |
| GSE102590 |
A Cdk9-PP1 switch regulates the elongation-termination transition of RNA polymerase II |
|
| Relations |
| BioSample |
SAMN08358592 |
| SRA |
SRX3557072 |
