New Resistance Gene Analog Polymorphism Primers Designed Based On Conserved Domain Sequences of Cereal Resistance Genes.

None of the previously designed resistance gene analog polymorphism (RGAP) primers was based on wheat and barley genes.  Here we report the development and use of RGAP markers using primers designed based on cloned wheat and barley genes for disease resistance.  The DNA sequence of the conserved motifs (P-loop, Kin2, NBS-B and GLPL) of Yr10, Lr1, Lr10, Lr21, Lr35, Pm3b and Mla1 and their start code flanking regions were used to design primers.  A total of 35 primers were designed. The length of the primer ranged from 15 to 18 nucleotides and the annealing temperature ranged from 38 to 54^oC.  PCR amplicons were resolved on 5.0-6.5% polyacrylamide gel or using an automatic Li-COR DNA analyzer. When various wheat genotypes were tested, the sizes of the amplicons ranged from 50 to 700 bp, and the average of the amplicon numbers was 74 (ranged from 42 to 131). Compared to previous primers designed based on resistance genes from other plant species, the bands produced by the new primers were more reproducible and robust. These primers were successfully used in mapping three wheat genes for stripe rust resistance in populations developed from wheat crosses AvS/PI 183527, AvS/PI 178759 and AvS/ PI 192252. The markers developed with these primers could be used in mapping resistance genes in wheat or barley, determining genetic diversity and identifying resistance genes.