|
| Status |
Public on Nov 02, 2015 |
| Title |
Drosophila_larvae_WT_CDK12_input_DNA_rep2 |
| Sample type |
SRA |
| |
|
| Source name |
third instar larvae
|
| Organism |
Drosophila melanogaster |
| Characteristics |
developmental stage: third instar
tissue: whole larvae
genotype: wild type
chip antibody: none
|
| Treatment protocol |
The w1118 strain was crossed with the actin-Gal4 strain as a wild type sample. For total larval chromatin preparation, approximately 3500 third instar larvae were collected, frozen in liquid nitrogen and stored at -80 °C.
|
| Growth protocol |
Larvae were cultured on standard medium (cornmeal/agar/sugar) in 18 degrees.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Using a ceramic mortar and pestle chilled with liquid N2, frozen larvae were ground to a powder and transferred to a homogenizer in ice-cold PBS. At room temperature, the homogenized cell suspensions were cross-linked with 1% formaldehyde for 20 min, quenched with 125 mM glycine for 5 min and filtered through Miracloth. Chromatin pellets were collected and resuspended in ice-cold RIPA buffer, followed by sonicating in a Branson Digital Sonifer with 25% output for 10 min (2s on, 4 s off). For immunoprecipitation, Protein A Sepharose™ Fast Flow (GE Healthcare 17-5280-01) was pre-blocked, 4μl CDK12 polyclonal antibody were used for each immunoprecipitation. The input DNA and the IP DNA were purified with a PCR purification Kit (Qiagen28104) and eluted in TE buffer.
DNA was end-repaired, adenylated, and ligated to TruSeq sequencing adaptors. DNA was purified by AMPure beads according to the DNA amount after each step. Library DNA was amplified by Phusion HF (NEB, Cat. M0530L). 2% agarose gel was used to select 200-500bp DNA fragments after PCR amplification.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 1500 |
| |
|
| Data processing |
The ChIP-seq reads were mapped to Drosophila melanogaster genome (dm3) using Bowtie2 with parameters -t -q -N 1 -L 25.
All unmapped reads, non-uniquely mapped reads and PCR duplicates were removed.
The occupancy profile of CDK12 was recorded into files in wiggle track format (WIG) for analyzing the data in UCSC Genome Browser
Genome_build: dm3
Supplementary_files_format_and_content: wig files for input in replicate 2.
|
| |
|
| Submission date |
Aug 30, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Li xia Pan |
| Organization name |
Tsinghua university
|
| Department |
School of medicine
|
| Lab |
C304
|
| Street address |
North Zhongguancun Street
|
| City |
Beijing |
| State/province |
Beijing |
| ZIP/Postal code |
100084 |
| Country |
China |
| |
|
| Platform ID |
GPL19951 |
| Series (1) |
| GSE63011 |
Heterochromatin remodeling by CDK12 contributes to learning in Drosophila |
|
| Relations |
| BioSample |
SAMN04018169 |
| SRA |
SRX1170640 |
