Sample information curated by ChIP-Atlas

Antigen

Antigen Class
Input control
Antigen
Input control

Cell type

Cell type Class
Larvae
Cell type
L3
NA
NA

Attributes by original data submitter

Sample

source_name
L3 larvae
tissue
whole larvae
strain
JA1507
Stage
L3
genotype/variation
lin-35 mutant
chip antibody
none

Sequenced DNA Library

library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
Worm_L3_extraction_vPK1. 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. Worm_chromatin_immunoprecipitation_vIL2. Appropriate amount of extract is incubated overnight with a proper amount of antibody (exceptional antibodies due to better results are incubated 2hrs). Afterwards, 40ul of equilibrated magnetic beads (either protein A or G, depending on antibody) are added and incubated for 2 hrs. Later, washes with FA, 500mM-salt FA, 1M salt FA, TEL, and TE buffer are performed and DNA is eluted in elution buffer (1% SDS in TE with 250 mM NaCl) two times with 57 ul volume each, at 65°C. Samples are treated with RNAse, proteinase K and then crosslinks are reversed overnight at 65°C. DNA is purified on Qiagen PCR purification columns, tested by q-PCR for ChIP quality, and stored at -20°C for future applications. DNA was incubated with an enzyme mix (Klenow, T4 DNA polymerase and T4 PNK) to ensure blunt ends and then with Exo(-) Klenow fragment in the presence of dATP to add adenosine at the 3' ends. The DNA fragments were ligated with appropriate adaptor (Illumina) and then amplified by PCR. The amplicon was loaded into an agarose gel, and size-selected DNA was recovered from the gel. Prepared sample are sequenced using Illumina GAIIx or HiSeq2000 at the High Throughput Sequencing Facility of University of North Carolina at Chapel Hill or Cambridge.

Sequencing Platform

instrument_model
Illumina Genome Analyzer IIx

ce11

Number of total reads
11911300
Reads aligned (%)
99.6
Duplicates removed (%)
6.9
Number of peaks
591 (qval < 1E-05)

ce10

Number of total reads
11911300
Reads aligned (%)
99.6
Duplicates removed (%)
6.9
Number of peaks
593 (qval < 1E-05)

Base call quality data from DBCLS SRA