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利用抗病基因的保守结构设计引物,从抗叶锈病近等基因系材料TcLr24中扩增出一条703bp的条带RGA1,通过与GenBank比对,选取与RGA1高度同源的若干条带,在它们共有的保守序列位置设计引物,利用cDNA末端快速扩增(RACE)技术扩增抗病同源基因cDNA全长。扩增到3条全长cDNA,经BLASTp比较,这些序列都含有NBS保守结构域和多个LRR结构域,与很多已知植物抗病基因的功能相应区域一致。对FRGA-1、FRGA-2和FRGA-3实时定量PCR分析,表明这3个基因在小麦叶片中都是组成型表达。本研究在小麦材料TcLr24中得到3条抗病基因同源cDNA全长,为研究小麦抗病基因奠定了基础。
A conservative 703bp band RGA1 was amplified from the near-isogenic line TcLr24 of leaf rust resistance by using the conserved structural design of the resistance gene. Several bands highly homologous to RGA1 were selected by comparison with GenBank, The conserved sequences were designed to amplify the full-length cDNA of disease-resistant homologous genes by using rapid amplification of cDNA ends (RACE). Three full-length cDNAs were amplified. Compared with BLASTp, these sequences contained NBS conserved domains and multiple LRR domains, which were consistent with the corresponding functional areas of many known plant disease resistance genes. Real-time quantitative PCR analysis of FRGA-1, FRGA-2 and FRGA-3 showed that all three genes were constitutively expressed in wheat leaves. In this study, we obtained the full-length cDNAs of three disease-resistance genes in wheat TcLr24, which laid the foundation for the study of disease-resistance genes in wheat.