de Wit Lab, Division Of Gene Regulation, Netherlands Cancer Institute
Description:
4C-Seq was performed as described under the Hi-C section. DNA extraction by phenol/chloroform, digested with a 2nd restriction enzyme (Csp6I). circular DNA molecules were created, then DNA was ligated under diluted conditions (10ml). DNA was precipitated with 1/10 volume 3M sodium acetate and 1 volume isopropanol. DNA was quantified using QUBIT (thermo fisher). 4C PCR was done using NEB-Next High-Fidelity (NEB,M0541) in 4 separate PCR reactions using 0.25-200ng per PCR