Molecular cloning and characterisation of a non-TIR-NBS-LRR type disease resistance gene analogue from sugarcane

Abstract

Disease resistance gene cloning is one of the main targets in modern sugarcane research. In this study, several primers were designed according to the conserved motifs in the NBS regions of known resistance genes to amplify sequences from cDNA of sugarcane variety NCo376 by polymerase chain reaction (PCR). In total, six NBS-LRR type resistance gene analogs (RGAs) were cloned, for which the deduced amino acids encoded all or parts of the internal motifs characteristic of the NBS-LRR R-gene class (Accession numbers: EF155648, EF155649, EF155650, EF155651, EF155652 and EF155653). One of them termed EF155653 had all the characteristics of NBS but had no significant similarity to any of the known genes, which suggested that it was possibly a partial sequence of a new NBS-LRR gene. Homology analysis was also conducted to evaluate the relationship between sugarcane RGAs and known plant R genes. Furthermore, all of them contained the residue W in LLVLDDVW/D motif, which confirmed that only non-TIR-NBS-LRR type resistance genes existed in sugarcane to some extent. Finally, the full-length cDNA of cRGA1 (Accession number: EF155648), termed SNLR gene, has been cloned and its expression profile under the treatment of Ustilago scitaminea, SA and H₂O₂ was investigated by real-time RT-PCR (Accession number: EF155654). The results showed that SNLR gene could be to some extent influenced by Ustilago scitaminea and SA, but not by H₂O₂.