Identification of PCR-based markers linked to the powdery-mildew-resistance gene Pl(1) from Malus robusta in cultivated apple

The availability of molecular markers linked to mildew resistance genes would enhance the efficiency of apple-breeding programmes. This investigation focuses on the identification of random amplified polymorphic DNA (RAPD) markers linked to the Pl(1) gene for mildew resistance, which has introgressed from Malus robusta into cultivated apples. The RAPD marker technique was combined with a modified 'bulked segregant analysis' mapping strategy. About 850 random decamer primers used as single primers or in combinations were tested by PCR analysis on the basis of resistant and susceptible DNA pools. Selected primers producing RAPD fragments were applied in an additional selection step to M. robusta and genotypes representing intermediate breeding stages of the breeding population 93/9, for which a 1:1 segregation could be observed for the resistance trait. Seven RAPD markers, all representing introgressed DNA sequences from M. robusta, were identified and arranged with the Pl(1) locus in a common linkage group. The two most tightly-linked RAPD markers, OPAT20(450) and OPD2(1000) were mapped with a genetic distance of 4.5 and 5 cM, respectively, from the Pl(1) gene. Both markers are suitable for marker-assisted selection in apple breeding. The polymorphic DNA fragment OPAT20(450) was cloned and sequenced, and longer primers for the generation of a sequence-characterized amplified region (SCAR) marker have been constructed; this marker was easier to score than the original RAPD marker.