HIGH OUTPUT GENETIC-MAPPING OF POLYPLOIDS USING PCR-GENERATED MARKERS

The polymerase chain reaction (PCR) with arbitrarily selected primers has been established as an efficient method to generate fingerprints that are useful in genetic mapping and genomic fingerprinting. To further increase the productivity of mapping and fingerprinting efforts, we have altered existing protocols to include the use of the Stoffel fragment, which is derived from genetically engineered Taq polymerase. We also optimized the thermal profile of the reaction to increase the number of useful primers. In mapping of the genome of Saccharum spontaneum 'SES 208', a polyploid wild relative of sugarcane, these modifications allowed for an increase of 30% in the number of loci screened per primer, and an 80% increase in the number of polymorphisms per primer. Furthermore, the enzyme cost per reaction was decreased approximately 1.6-fold. Finally, there was an increase from about 70% to about 97% in the number of primers that were useful (i.e., gave a reproducible fingerprint) using our protocol. We have placed some of these markers into linkage groups.