Catch-linker + PCR labeling: a simple method to generate fluorescence in situ hybridization probes from yeast artificial chromosomes.

Abstract

A simple and efficient method to generate hapten-labeled DNA fragments from a trace amount of YAC DNA isolated by PFGE is described. After agarase digestion of the gel slice containing the resolved YAC recombinant, the purified DNA is digested with Sau3Al and a compatible CL oligonucleotide duplex ligated on. A probe is generated by PCR amplification using a primer complementary to the CL with a single biotin moiety incorporated at the 5' end. When used as a FISH probe, this material yields mapping results superior to Alu-PCR or whole YAC labeling methods and allows sensitive detection of chimerism.