Use of polymerase chain reaction to screen phage libraries.

ABSTRACT

Isolating a clone from a cDNA or genomic library often involves screening the library by several rounds of plating and filter hybridization. This is not only laborious and time-consuming, but also is prone to artifacts such as false positives commonly encountered in filter hybridization. These problems can be alleviated by using polymerase chain reaction (PCR) in the early rounds of screening prior to conventional filter hybridization. The advantages of PCR screening are threefold (1) Positive clones are identified by DNA bands of correct sizes in gel, thus avoiding the confusion from false positive spots in filter hybridization; (2) it saves time, especially in initial rounds of screening; and (3) screening of multiple genes can be performed in the same PCR by using appropriate primers for these genes. After the complexity of the phage pool is reduced and the existence of true positives in the pool confirmed, individual clones can be isolated by conventional methods.