GENERATION OF POLYMORPHIC MARKERS TIGHTLY LINKED TO THE THYMUS ENLARGEMENT LOCI BY PHENOTYPE-DIRECTED REPRESENTATIONAL DIFFERENCE ANALYSIS

To obtain genetic markers linked to a specific genetic locust genomic subtraction with a DNA pool of backcross or F-2 intercross animals wit h a specific genotype at the locus is known to be effective. To determ ine whether the pooling strategy is also effective for isolation of ge netic markers linked to a quantitative phenotype that can potentially be controlled by multiple genetic loci, we tested the ability of repre sentational difference analysis (RDA) to isolate genetic markers linke d to the thymus enlargement observed in the BUF/Mna (BUF) rat. This is known to be controlled by single major and minor genes, Ten1 and Ten2 , on Chromosomes (Chrs) 1 and 13, respectively, both of which have dos e effects on the normal WKY/Ncj (WKY) allele. DNA from an inbred WKY r at was used as the tester, and the driver was prepared from a DNA pool of 12 (WKY x BUF)F-1 x BUF backcross rats with high thymus ratios (th ymus weight/body weight), expected to have dominance of the BUF allele in the responsible loci. By two RDA series with the restriction enzym es BglII and BamHI, respectively, 28 polymorphic markers were isolated , and 8 of them were shown to be linked to Ten1, and one to Ten2. One of the 8 markers linked to Ten1 demonstrated no recombination in 18 ra ts with high thymus ratios. RDA with a DNA pool based on a quantitativ e phenotype (phenotype-directed RDA) can thus be considered an efficie nt approach for direct isolation of polymorphic markers linked to a qu antitative trait.