东乡野生稻NBS类抗病基因同源片段的克隆及序列分析

Cloning and Sequence Analysis of NBS Resistance Gene Analogs from Dongxiang Oryza rufipogon Griff

根据已知的NBS-LRR类抗病基因结构中氨基酸的保守区域,设计简并引物,通过RT-PCR扩增及克隆,从东乡野生稻(Dongxiang Oryza rufipogon Griff.)中共获得7个新的NBS-LRR类抗病基因同源片段。所有7个抗病基因同源序列均含有NBS-LRR类抗病基因的保守序列,如P-loop、Kinase 2、Kinase 3a以及跨膜区域等。通过氨基酸同源性比较和分析,发现克隆到的抗病基因同源序列与已克隆的水稻白叶枯抗病基因Xa1的相应氨基酸序列存在约40%的同源性,其中的3个NBS抗病基因同源序列还与小麦抗叶锈病基因Lr1存在40%的同源性。

Degenerate oligonucleotide primers were designed on the basis of nucleotide-binding site（NBS）motifs conserved among the reported disease resistance genes.By reverse transcription polymerase chain reaction（RT-PCR）,seven new NBS-LRR disease resistance gene analogues have been obtained and cloned from Dongxiang Oryza rufipogon Griff.Sequencing of the DNA fragments indicated that all of NBS resistance gene analogues contained the conserved motifs of NBS-LRR type resistance genes,such as P-loop（Kinase 1a）,Kinase 2,Kinase 3a and transmembrance domain.By comparison analysis of amino acid sequences,it was found that the obtained disease resistance gene analogues have about 40%identity with the reported bacterial blight-resistance gene Xa1.Three of them have about 40%identity with the reported wheat leaf rust resistance gene Lr1.