大豆抗病基因同源序列的克隆与分析

Cloning and Analyzing of Resistance Gene Analogs in Soybean ZDD2315

本研究根据已知抗病基因的NBS保守序列区设计 4对简并引物和 1对特异引物 ,以大豆农家种兴县灰布支黑豆为材料 ,应用PCR方法获得了 11条来自基因组DNA的RGA序列和 2条来自cDNA的RGA序列 ,序列长度在 5 0 0 - 6 33bp之间 ,其中 8条来自基因组DNA和 2条来自cDNA的RGA序列已在GeneBank登录 (登录号为 :AF30 5 388- 30 5 392 ,AY0 0 8380 - 0 0 8382 ,AY0 4 886 3-AY0 4 886 4)。 13条序列都不同程度的含有NBS保守区的P -环 (GGVGKTT)、kinase - 2 (VLDD)、kinase - 3(GSRII)及跨膜区GLPL等特征序列结构 ,由此推导出的氨基酸序列同已知抗病基因L6、RPM 1、SRPS2、N编码的氨基酸序列表现出从 2 5 % - 42 %的同源性。本研究克隆的RGA序列根据其相似性可分为 4组 ,与已发表的大豆抗病类似基因 (RLG)具有较高的相似性。

Based on the NBS conserved sequences of known resistance genes, a pair of specific primer and four pairs of degenerate primers were designed to be used for amplifying soybean resistance gene analogs (RGAs). In this study, eleven RGAs from genomic DNA and two RGAs from cDNA were obtained in soybean ZDD2315 (Glycine max), which of ten sequences have been registered in the Genebank,(Accession number: AF305388-92, AY008380-82, AY048863-64). Sequence analysis of cloned RGAs indicated that the size of cloned RGAs are between 500bp to 633bp in length and four motifs of NBS conserved regions exist in the sequenced RGAs, which including P-loop (GGVGKTT), kinase-2(VLDD), kinase-3(GSRII) and membrane spanning region (GLPL). The amino acid sequences deduced from the sequenced RGAs show 25%-42% homology with amino acid sequences of L6?RPM1?SRPS2?N genes. These cloned RGAs could be divided into four gounps and have more similarity compared with the published RLGs(Resistance Like Gene).