Heterologous Expression and Characterization ofToxA1haplotype from India and its interaction withTsn1for Spot Blotch Susceptibility in spring wheat

Abstract Background ToxA , a necrotrophic effector protein, is present in the genome of fungal species like Parastagnospora nodorum , Pyrenophora tritici-repentis and Bipolaris sorokiniana. Tsn1 is the sensitivity gene in the host whose presence indicates more susceptibility to ToxA carrying pathogen, and ToxA - Tsn1 interaction follows an inverse gene-for-gene relationship. Methods and Results The present study involved cloning and expressing the ToxA1 haplotype from B. sorokiniana . It was found that the amplicon exhibited an expected product size of 471 bp. Sequence analysis of the ToxA1 nucleotide sequence revealed the highest identity, 99.79%, with P. tritici-repentis . The protein expression analysis showed peak expression at 16.5kDa. Phylogenetic analysis of the ToxA1 sequence from all the Bipolaris isolates formed an independent clade along with P. tritici-repentis and diverged from P. nodorum . ToxA-Tsn1 interaction was studied in 18 wheat genotypes (11 Tsn1 and 7 tsn1 ) at both seedling and adult stages, validating the inverse gene-for-gene relationship, as the toxin activity was highest in the K68 genotype ( Tsn1 ) and lowest in WAMI280 ( tsn1 ). Conclusion The study indicates that the haplotype ToxA1 is prevailing in the Indian population of B. sorokiniana . It would be desirable for wheat breeders to select genotypes with tsn1 locus for making wheat resistant to spot blotch.