Abstract
The pathogenic fungus, Bipolaris sorokiniana, that causes spot blotch (SB) disease of wheat, is a major production constraint in the Eastern Gangetic Plains of South Asia and other warm, humid regions of the world. A recombinant inbred line population was developed and phenotyped at three SB-prone locations in India. The single nucleotide polymorphism (SNP) for SB resistance was identified using a bulked segregant RNA-Seq-based approach, referred to as "BSR-Seq." Transcriptome sequencing of the resistant parent (YS#24), the susceptible parent (YS#58), and their resistant and susceptible bulks yielded a total of 429.67 million raw reads. The bulk frequency ratio (BFR) of SNPs between the resistant and susceptible bulks was estimated, and selection of SNPs linked to resistance was done using sixfold enrichments in the corresponding bulks (BFR >6). With additional filtering criteria, the number of transcripts was further reduced to 506 with 1055 putative polymorphic SNPs distributed on 21 chromosomes of wheat. Based on SNP enrichment on chromosomal loci, five transcripts were found to be associated with SB resistance. Among the five SB resistance-associated transcripts, four were distributed on the 5B chromosome with putative 52 SNPs, whereas one transcript with eight SNPs was present on chromosome 3B. The SNPs linked to the trait were exposed to a tetra-primer ARMS-PCR assay, and an SNP-based allele-specific marker was identified for SB resistance. The in silico study of these five transcripts showed homology with pathogenesis-related genes; the metabolic pathway also exhibits similar results, suggesting their role in the plant defense mechanism.