En masse cloning of expressed disease resistance genes of wheat (Triticum aestivum) using RNA differential display via degenerate primers and data mining methods

Genetic control via manipulating resistance genes is the most effective and economical method of controlling plant pests using two conventional approaches, map-based cloning and transposon tagging. The objective of our study was to identify expressed resistance genes and physical localize them. Using modified RNA fingerprinting and data mining approaches, we totally identified 220 expressed resistance gene candidates out of the 728 analyzed. Irrespective of the cloning methods used, 184 out of 220 (36 unknown were eliminated) expressed resistance gene candidates were attempted to map physically using aneuploids and 339 deletion lines. Of the 184 sequences, 87 were NB/LRR type, 16 were receptor-like kinase (Xa21 type), 13 were protein kinase (Pto type), 7 were Hm1, 2 were Hs1pro-1, 17 were pathogen related and 42 were novel nucleotide bind ing/kinase-encoding type of sequences. Physical mapping localized 310 resistance loci in wheat genome detected by the 121 candidate resistance gene sequences. We combined the physical mapping information from the three homeologous groups to generate high-resolution consensus physical maps as the gene distribution and synteny is conserved among three wheat genomes. A total of 121 probe detected 151 loci, of which 143 were clustered in 26 smaller chromosomal regions encompassing ∼16% of the genome. Five major resistance gene clusters were observed to contain loci corresponding to 67 sequences (55% of the total). Wheat genome contains 269 morphologically characterized resistance genes. Two hundred twenty-nine of them were genetically studied. Construction of consensus genetic maps using individual 137 genetic linkage maps allowed us to localize 110 (80 single gene inherited and 30 QTL type) morphologically characterized wheat resistance genes on the consensus genetic maps. Comparison of consensus genetic maps with consensus physical maps, 80 single-gene-inherited and 10 QTL-type wheat resistance genes were physically located in 20 small physical chromosomal regions. About 93% (85 of 90) of the wheat resistance genes were localized into 18 smaller regions where 131 of the candidate resistance gene loci mapped. About 75% of both the morphologically characterized resistance genes and the identified candidate sequences mapped to the distal 20% of the chromosomal regions. (Abstract shortened by UMI.