|
| Status |
Public on May 02, 2011 |
| Title |
seq-SDQ4663_RPC1_N2_MXemb_1_A_EE8 |
| Sample type |
SRA |
| |
|
| Source name |
whole body
|
| Organism |
Caenorhabditis elegans |
| Characteristics |
tissue: whole body
strain: N2
Stage: L3
chip antibody: SDQ4663_RPC1
chip antibody supplier: SDI
|
| Treatment protocol |
Embryos were resuspended in FA buffer (50 mM HEPES/KOH pH 7.5, 1 mM EDTA, 1% Triton X-100, 0.1 % sodium deoxycholate; 150 mM NaCl) + protease inhibitors (Calbiochem Cat# 539131). Using a Branson sonifier microtip, samples were sonicated on ice at the following settings: 35% amplitude, 0.9 sec on, 0.1 sec off, 12 pulses, 10 times. Cell debris was removed by centrifuging at 13,000 g for 15 minutes at 4?C and taking the supernatant. Protein concentration was determined by Bradford Assay and extracts were aliquoted at stored at -80C. http://wiki.modencode.org/project/index.php/Worm_embryo_extraction:JL:2
|
| Growth protocol |
Embryos were prepared by bleaching from gravid N2 adults grown in standard S-basal media liquid culture. Live embryos were cross-linked in M9 + 2% formaldehyde for 30 minutes at room temperature followed by quenching with 125mM glycine for 5 minutes. Embryos were then washed twice with M9 Buffer and once by FA buffer (50 mM HEPES/KOH pH 7.5, 1 mM EDTA, 1% Triton X-100, 0.1 % sodium deoxycholate; 150 mM NaCl). Pellets were frozen at -80C. http://wiki.modencode.org/project/index.php/Worm_embryo_growth_and_harvest:JL:5
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Libraries were prepared according to Illumina's instructions for "Preparing Samples for ChIP Sequencing of DNA", with some variation. DNA was end-repaired using T4 DNA polymerase, Klenow DNA polymerase and T4 DNA polymerase. A's were then added to the 3' ends of the fragments using Kelow fragment (3' 5' exo minus). Paired-end adapters were then ligated onto the ends of the fragments. DNA library was size selected by gel purification, fragments of 250-500 bp were band isolated from an agarose gel. DNA was PCR amplified for 18 cycles. The Bioanalyzer was used to verify fragment purification and quantify DNA.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina Genome Analyzer II |
| |
|
| Description |
input control: GSM706160
Chromatin IP against RPC1. http://wiki.modencode.org/project/index.php/Worm_chromatin_immunoprecipitation:JL:KI3
|
| Data processing |
BWA is a fast light-weighted tool that aligns relatively short sequences to a sequence database. The algorithms is based on Burrows-Wheeler Transform (BWT). See the link @ http://bio-bwa.sourceforge.net/. It runs as DEFAULT for both bwa aln and bwa samse.
ChIP-Seq MACS Peak-calling:JL:1. Peak calls were determined by using Model-based Analysis of ChIP-Seq (MACS) software. MACS was supplied with the following parameters: tagSize = 25 mfold = 10,30 genomeSize = 90000000 bandwidth = 300 pvalue = 1e-5
Processed data file build: ce6
|
| |
|
| Submission date |
Apr 21, 2011 |
| Last update date |
May 15, 2019 |
| Contact name |
Tristan De Buysscher |
| Organization name |
University of North Carolina at Chapel Hill
|
| Department |
Biology
|
| Lab |
Lieb
|
| Street address |
CB# 3280, Coker Hall
|
| City |
Chapel Hill |
| State/province |
NC |
| ZIP/Postal code |
27599-3280 |
| Country |
USA |
| |
|
| Platform ID |
GPL9269 |
| Series (1) |
|
| Relations |
| SRA |
SRX059271 |
| BioSample |
SAMN00261290 |
| Named Annotation |
GSM713174_seq-SDQ4663_RPC1_N2_MXemb_1_A_EE8_macs14_MACS_wiggle_cat.wig.gz |
