Screening a BAC library with nonradioactive overlapping oligonucleotide (overgo) probes.

Comparative mapping has been the primary approach for analyzing genomes of divergent species because gene linkage is often conserved over evolutionary time, a phenomenon known as synteny. Comparisons are based on computational detection of orthologous sequences between species. Small overlapping oligonucleotide (overgo) probes, consisting of two self-annealing oligonucleotides at the 3'-end, are filled in with labeled deoxynucleotides. Overgo probes are designed against highly conserved regions between two genomes, at regular intervals (30-40 kbp). All methodologies reported to date have used radioactive dATP and dCTP to label overgo probes. We report a nonradioactive labeling protocol to produce digoxigenin-labeled overgo probes for high-throughput screening of the Actinidia chinensis bacterial artificial chromosome library.