Plant, Animal & Microbe Genomes X Conference

January 12-16, 2002
Town & Country Convention Center
San Diego, CA

Poster: Large Insert Libraries
            

MAP-BASED CLONING OF LATE BLIGHT RESISTANCE ALLELES FROM SOLANUM BULBOCASTANUM: BAC CONTIG DEVELOPMENT AND SEQUENCE ANALYSIS.

¹ Department of Plant Pathology, University of Wisconsin, 1630 Linden Dr., Madison WI 53706

² Department of Horticulture, University of Wisconsin, 1575 Linden Dr., Madison WI 53706

We have previously mapped genes conditioning late blight resistance to the long arm of Solanum bulbocastanum chromosome 8. FISH analyses indicate that the physical distance separating the nearest flanking markers is slightly less than 1MB. We have constructed a partial BAC contig and saturated the resistance region using markers derived from BAC end sequences, localizing the genes responsible for the resistant phenotype to within 55 kb. The S. bulbocastanum plant used to construct the BAC library is heterozygous for alleles conditioning resistance. Using PCR-based markers derived from BAC ends, we have determined homolog origin for each BAC clone encompassing the 55 kb resistance region; each of 12 clones originated from the susceptible homolog. A single BAC clone of approximately 165 kb was sequenced in its entirety. This clone, derived from the susceptible homolog, contains the entire genetically defined resistance region. The results of detailed sequence analyses will be discussed. Using a series of overlapping long-range PCR primers designed from the susceptible homolog BAC, approximately 70 kb encompassing the genetically defined resistance region was amplified from genomic DNA. Cloning and subsequent marker analyses revealed that the PCR products are from both homologs. Sequence analysis and large insert transformation experiments of the resistant homolog long-range PCR products are underway.