利用RGA-PCR方法进行水稻抗瘟基因分子标记

RGA-PCR Marking of Resistance Genes to Rice Blast

用28对RGA引物对LTH近等基因系品种进行PCR扩增,其中11对引物扩增出特异性条带。将扩增到的33个特异性片段回收并进行重扩增,有11个片段产生单一条带。选择2个片段HS-1和HS-19进行探针标记。经Southern杂交发现,探针HS-1在含有Pi-ta2抗瘟基因品种F-128-1、NO4中有特异杂交信号,表明该片段可能与抗瘟基因Pi-ta2连锁或是其一部分。对特异性片段HS-1进行克隆、测序,全长为478bp,与Mago等从水稻中克隆的一个抗病基因同源序列RGA29有95%同源性。

28 primer pairs designed according to plant resistance gene analog were used to amplify rice LTH NILs. Of them, 11 primer pairs gave peculiar bands. 33 peculiar bands were retrieved and re-amplified, and 11 single bands were acquired. Fragment HS-1 and HS-19 with the size of 480 and 490 bp were respectively selected as probes. Southern hybridization showed that HS-1 fragment could detect peculiar signals in rice varieties with resistance gene Pi-ta^2, which revealed that this sequence could link with Pi-ta^2gene or be a part of the gene. HS-1 fragment was cloned and sequenced, and it had 478 bp. The sequence was 95% homogenous with RGA29, a resistance gene analog cloned from rice by Mago et al.