A SPOT CLONING METHOD FOR RESTRICTION LANDMARK GENOMIC SCANNING

We introduce two new methods for target cloning of DNA fragments corre sponding to spots on the two-dimensional profile of restriction landma rk genomic scanning (RLGS). One is a restriction trapper-based method and the other is a polymerase chain reaction (PCR) mediated method. Bo th are designed to select the target DNA fragments from a large amount of unlabeled background DNA fragments in the RLGS gel which produce b ackground clones. The restriction trapper method is simple, with a clo ning efficiency that is not biased by the length of the target DNA nor by its GC content. On the other hand, the PCR-mediated method is effi cient for cloning DNA fragments from a small amount of starting materi als. These methods provide us with powerful tools for isolating DNA cl ones identified by the RLGS system as interesting spots. This paper re ports the precise protocols of these methods and discusses their appli cation and usefulness.