Library |
Library name |
Construction protocol |
Strategy |
Source |
Selection |
Layout |
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ChIP was performed according to Agilent Mammalian ChIP-on-chip manual. Briefly, 1x108 ES cells were fixed with 1% formaldehyde for 10 minutes at room temperature. Then the reactions were stopped by 0.125M glycine for 5 min. Chromatin were sonicated to an average of 200-500bp (ChIP-Seq) or 500bp-1000bp (ChIP-qPCR) using Covaris Sonication System (USA) according to the manual. Then 1% Triton X-100 was added to the sonicated chromatin solutions to a final 0.1% Triton X-100. After centrifuge, 50 ul of supernatant were saved as input. The rest of chromatin solution was incubated with Dynabeads previously coupled with ChIP grade antibodies overnight at 4℃ with rotation. After extensive washes with washing buffer, the complexes were reverse cross-linked overnight at 65℃. DNAs were extracted by hydroxybenzene-chloroform-Isoamyl alcohol and purified by Phase Lock Gel (Tiangen, China). The ChIPed DNA were used for library constructions and deep sequencing by Shenzhen BGI (Wuhan, China). For ChIP-PCR, ChIPed DNA were quantified by quantitative real-time PCR(qPCR). The enrichment was calculated relative to the amount of input chromatin. |
ChIP-Seq |
GENOMIC |
unspecified |
SINGLE
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