|
| Status |
Public on May 02, 2011 |
| Title |
seq-BH00004_LIN54_N2_L3_1 |
| Sample type |
SRA |
| |
|
| Source name |
whole body
|
| Organism |
Caenorhabditis elegans |
| Characteristics |
tissue: whole body
strain: N2
Stage: L3
chip antibody: BH00004_LIN54
chip antibody supplier: Bob Horvitz
|
| Treatment protocol |
Worms are frozen, ground, and crosslinked for 10 minutes in 1% formaldehyde. Formaldehyde is quenched and cross-linked tissue washed, then resuspended in FA buffer and subjected to sonication in Bioruptor (14 pulses of 30 seconds with 1 minute rests in between). Extracts are then spun down and soluble fraction is stored for quality tests and future ChIP. http://wiki.modencode.org/project/index.php/Worm_L3_extraction:JL:PK1
|
| Growth protocol |
About 2-7 million of worms are bleached and then hatched in M9 for 24-42 hrs. About 100 embryos are seeded onto the plate to test for contamination and hatching efficiency. Remaining hatched L1 larvae are inoculated in a proper volume of liquid culture. Next day when larvae reach the L3 stage they are cleaned by M9 washes and sucrose gradient and collected by freezing in liquid nitrogen. Just before collection DIC pictures are taken and about 50ul of worms are stained for DAPI to assess the stage. http://wiki.modencode.org/project/index.php/Worm_L3_growth_and_harvest:JL:PK1
|
| 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: GSM706161
Chromatin IP against LIN54. http://wiki.modencode.org/project/index.php/Worm_chromatin_immunoprecipitation:JL:IL2
|
| 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 |
SRX059232 |
| BioSample |
SAMN00261251 |
| Named Annotation |
GSM712713_seq-BH00004_LIN54_N2_L3_1_macs14_MACS_wiggle_cat.wig.gz |
