小麦NBS-LRR类抗病基因同源cDNA序列的克隆与表达分析  

Cloning and expression analysis of a NBS-LRR resistance gene homology cDNA sequence from wheat

作　　者：任晓娣[1] 刘彦慧[1] 李建嫄[1] 张娜[1] 彭巧慧 杨文香[1] 刘大群[1] 

机构地区：[1]河北农业大学植物病理学系/河北省农作物病虫害生物防治工程技术研究中心/国家北方山区农业工程技术研究中心,河北保定071001 [2]河北省农科院,河北石家庄050011

出　　处：《河北农业大学学报》2013年第2期69-74,79,共7页Journal of Hebei Agricultural University

基　　金：河北省自然科学基金项目(C2006000438);国家重点基础研究发展计划(2013CB127700)

摘　　要：为获得小麦抗叶锈病相关基因,以小麦近等基因系TcLr19所构建的非亲和cDNA文库中获得的EST序列(Contig 914)为靶序列,用RT-PCR方法和cDNA末端快速扩增技术,分离克隆到片段为3 042bp的全长cDNA序列。序列分析表明该序列符合典型单子叶植物的CC-NBS-LRR结构模式,命名为TaNLR。该基因包含一个完整的2 739bp的开放阅读框(ORF),具有连续的Poly A尾和典型的加尾信号AATTAA。ProtParam程序预测表明该基因编码912个氨基酸。发育树分析显示该氨基酸序列与大麦的NBS-LRR类抗病基因蛋白同源性最高达89%。荧光定量PCR分析表明,在小麦与叶锈菌互作中,TaNLR基因受叶锈菌诱导下调表达。本研究在TcLr19小麦中成功获得了抗病同源基因,这为明确NBS-LRR在小麦抗叶锈病中的作用奠定了基础。Reverse transcription-polymerase chain reaction （RT-PCR） and rapid amplification of cDNA ends （RACE） were applied in cloning disease resistance-associated gene with ESTs （Contig 914） as target sequence came from incompatible interaction cDNA library of TcLr19 to obtain wheat leaf rust resistance-associated gene. The full length cDNA of the aimed gene is 3 042 bp and containing an open reading frame of 2 739 bp. The sequence analysis showed that the gene belonged to the CC-NBS-LRR type gene, temporarily designated as TaNLR. TaNLR has continuous poly A tail and a typical poly adenylation signal. The ProtParam program puta tively predicted that this gene encoded a protein of 912 amino acids. The phylogenetic tree anal- ysis showed 89% identity of the protein encoded by TaNLR with that of Hordeum vulgate. Real-time PCR analysis revealed that TaNLR gene was induced and down-regulated expression during interaction between P. triticina and TcLrl9. The resistance homology sequence was successfully obtained in TcLr19, which laid the foundation for understanding the function of NBS-LRR to wheat leaf rust resistance.

关 键 词：小麦 核苷酸结合位点(NBS) 富含亮氨酸重复(LRR) 抗病基因同源序列(RGAs) cDNA末端快速扩增技术(RACE) 实时定量PCR(real-time PCR) 

分 类 号：S435.121.43[农业科学—农业昆虫与害虫防治]