CANDIDATE RESISTANCE GENES FROM AN EST DATABASE PROVE A RICH SOURCE OF MARKERS FOR MAJOR GENES CONFERRING RESISTANCE TO IMPORTANT APPLE PESTS AND DISEASES

An apple EST database containing over 100,000 sequences from 44 cDNA libraries representing several cultivars and a range of tissues has provided an excellent resource of genetic markers linked to previously identified resistances to apple scab, powdery mildew and woolly apple aphid.
Candidate ESTs were mined from this database on the basis of homology to genes from 5 resistance (R) gene classes and screened as RFLP probes over Southern blots of DNA from seedling populations segregating for 5 selected resistances to pest or pathogen infection.
We have used this targeted strategy to obtain efficiently a set of additional markers for a number of resistance genes that are candidates for marker assisted selection in our apple breeding program, namely the V_f, Pl₂, Pl_MIS, and Er₃ genes.
Steps used in our targeted strategy include Southern analysis with mini-population blots to identify possible linkages, screening of enlarged populations to confirm linkage, and then conversion of the more closely linked markers to PCR based markers (SCAR, SNP) to enable high throughput population screens.
We have established that (as anticipated) candidate ESTs used as RFLP probes do not need to be derived from varieties that carry recognized R genes.
Linkage analysis suggests that these ESTs often cluster around the R gene in a region of several centiMorgans (cM). This region often includes clusters that can contain candidates from more than one R gene class.
Some R genes derived from distinct backgrounds have genetic markers in common.