Nucleotide binding site/leucine-rich repeats, Pto-like and receptor-like kinases related to disease resistance in grapevine

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Abstract
Nucleotide Binding Site/Leucine-Rich Repeat (NBS-LRR) and Serine/Threonine Kinase (STK) genes are two of the known classes of resistance (R-) genes in plants, and occur in large multigene families. Systematic identification of genes for NBS-LRRs and STKs provides a means of access to genomic regions that may be involved in disease resistance. Here we present a picture of these two families of R-gene analogs (RGAs) in grape with the aim of developing a set of resistance-related sequence-tagged-site (STS) markers. One hundred and three NBS-LRR sequences were isolated. They included members of the CC (coiled-coil) and TIR (Toll-interleukin receptor) sub-classes. A comparative analysis with other angiosperm NBSs is provided. Fifty-three genes for receptor-like kinases (RLKs) with serine/threonine specificity were identified. RLK sequences formed a putative monophyletic group within the kinase superfamily. They were similar to both cytoplasmic RLKs, such as Pto, and RLKs with LRR, S-locus, lectin-like and thaumatin-like extracellular binding-domains. The latter resembled the products of the R-related genes Xa21, FLS2, Rlk10, SFR2, and PR5K. Forty-five reference RGAs were converted into STSs by using appropriately designed specific primers. RGA-STSs were present in diverse grape genotypes, and >85% of the primers were capable of amplifying the STSs across the taxa Vitis and Muscadinia. DNA sequence polymorphism among these RGAs was assessed by SSCP (single-strand conformation polymorphism) analysis in over 20 Vitis spp. Finally, 45 universal primers for grape RGAs are proposed that should permit tagging of R-related regions in any grape genome.