2.6 Marker designing and Genotyping of F2 population
For genotyping of DOC3.2 locus in F2 population (derived from crosses between B1K-05-07 and B1K-50-04, we identified sequence diversity using Nanopore data covering region of DOC3.2(Chr3H_67433882-Chr3H_ 67434491). We found SNP marker Chr3H_67434330 have C to T in B1K-50-04 and B1K-05-07, respectively, that resides within the TaqI recognition. For genotyping of F2 population, we isolated leaf DNA using with CTAB method, PCR amplified using PM.22-0190 and PM.22-0191 primers (Table S1 ) followed by restriction digestion using TaqI restriction enzyme to distinguish between three genotypes.