5C Analysis of 3C, ChIP-Loop, and Control Libraries

Abstract

This protocol describes the 5C method of detecting ligation products in 3C, ChIP-loop, or control libraries. The method starts with a purified 3C library, a purified ChIP-loop library, or a purified control library. Before starting the protocol, assess the quality of the library that will be used by performing semiquantitative polymerase chain reaction (PCR). Ligation products should be readily detectable and nearby restriction fragments should be ligated more frequently than distant fragments. The design of the 5C primers and methods to determine interaction frequencies are also described in this protocol.