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.
Hilario
Elena
E
Gene Technologies, HortResearch Ltd., Auckland, New Zealand.
Bennell
Tiffany F
TF
Rikkerink
Erik
E
D029042:Actinidia / Q000235:genetics; D022202:Chromosomes, Artificial, Bacterial / Q000235:genetics*; D018744:DNA, Plant / Q000235:genetics / Q000302:isolation & purification; D004076:Digoxigenin; D018745:Genome, Plant; D015698:Genomic Library; D049449:Luminescence; D015336:Molecular Probe Techniques; D009693:Nucleic Acid Hybridization; D015345:Oligonucleotide Probes
D018744:DNA, Plant; D015345:Oligonucleotide Probes; D004076:Digoxigenin