DNA marker development by the allele-specific detection of powdery mildew resistance loci derived from Japanese domestic tobacco cultivar ‘Kokubu’

Japanese domestic tobacco (Nicotiana tabacum L.) cultivar ‘Kokubu’ shows high powdery mildew resistance controlled by recessive alleles at two loci, and these alleles have been widely used as a resource for powdery mildew resistance in tobacco breeding. However, the introduction of this trait by conventional breeding takes much work because of the requirement for test crosses with the parental strains and inoculation tests using active fungi to confirm the introduction of two recessive alleles during back-crossing. Recently, we found that powdery mildew resistance in ‘Kokubu’ is caused by splice site mutations of two MILDEW LOCUS O genes, NtMLO1 and NtMLO2. Here, we report DNA markers that detect mutations of the NtMLO1/2 genes based on the cleaved amplified polymorphic sequence (CAPS) or allele-specific polymerase chain reaction (AS-PCR) methods. These markers can be used as co-dominant markers that detect heterozygotes of the NtMLO genes at the seedling stage in back-crossed progenies, and will contribute to the simplification of breeding.