Plant Genome III Conference Plant Genome III Conference
A targeted mapping strategy using representational difference analysis (RDA) (Lisitsyn et al., 1993) was employed to isolate new RFLP probes for the long arm of chromosome 6 in rye (6RL), which carries a gene for resistance to Hessian fly larvae. Fragments from the 6RL arm were specifically isolated using a Chinese Spring (CS) wheat-rye ditelosomic addition line (CSDT6RL) as the tester and CS and/or CS4R as the driver for the genomic subtraction. Three RDA experiments were performed with Bam HI amplicons, two of which were successful in producing low copy dories. All low copy clones were confirmed to have originated from 6RL, indicating substantial enrichment for target sequences. However, the ability of the technique to achieve significant enrichment was found to be dependent on 1) the condition of the starting plant material (i.e. presence of contaminating pathogens), 2) the order the three primer sets were used in, and 3) the genomic complexity of the amplicons. Two mapping populations, both of which are derived from a cross between two similar wheat-rye translocation lines, were used to map five RDA probes as well as five wheat probes. One of the populations was prescreened for recombinants by C-banding analysis. Fifteen loci including seven RDA markers, were placed on a map of the distal half of 6RL. Four ofthe RDA markers are within 1O-12cM ofthe Hessian fly resistance gene, which was located by mapping and C-banding analysis to approximately the terminal 1% of the arm.