title: HNF6 ChIP-seq in mouse
design: HNF6 ChIP-seq in mouse
library_construction_protocol: The animal livers were crosslinked with formaldehyde treatment and chromatin fragmented to an average of 300 bp by sonication. Chromatin from an mass equivalent of 1/4 mouse liver was used for each ChIP experiment. Solexa libraries were prepared following the instructions of Illumina (Sample preparation for genomic DNA version 2.2) with the following modifications. The ChIP-enriched DNA and input DNA were not further fragmented. After end-repair and addition of an A base to the 3 ends, the adapters were ligated to the ends of the DNA Fragments using 2 ml of fourtyfold diluted Adapter oligo mix in a total reaction volume of 25 ml. Between these steps the DNA was purified using the DNA Clean-and-Concentrator-5 kit (Zymo Research). Subsequently the DNA was amplified by 18 cycles of PCR purified with QIAquick PCR purification Kit and eluted with 33.5 ml of 10 mM tris buffer at pH7.0. The PCR-product was sized fractionated on 2 percent agarose gel and a gel slice containing the 200-300 bp fragments was excised. Hepatocytes were prepared by direct perfusion of the liver with buffered salt solution followed by percent formaldehyde. After 10 minutes the tissue was removed and diced in 500 mM glycine buffer to neutralize the formaldehyde. After homogenization the hepatocytes were rinsed with PBS.
identifiers.namespace: CRUK-CRI
identifiers.id: E-MTAB-438:do40HNF6SAN03