不结球白菜抗病基因同源序列的克隆及分析.pdfVIP

中国农业科学 2006,39(12):2621-2626 Scientia Agricultura Sinica 不结球白菜抗病基因同源序列的克隆及分析 1,2 1 2 2 2 王彦华 ，侯喜林 ，申书兴 ，陈雪平 ，王 梅 1 2 （南京农业大学作物遗传与种质创新国家重点实验室，南京 210095 ； 河北农业大学园艺学院，保定 071001 ） 摘要: 【目的】利用同源序列法分离不结球白菜抗病基因同源序列。【方法】根据植物抗病基因TIR-NBS-LRR 保守区设计简并引物，对不结球白菜基因组DNA及cDNA 进行PCR扩增。【结果】获得了10个具有通读氨基酸序列 的片段。同源性比较发现，该10个片段均属于TIR-NBS-LRR类抗病基因同源序列（RGA），与已知R基因相应区段 的氨基酸序列一致性为 50%～66%。与已知抗病基因聚类分析结果显示，该 10 个 RGA 序列可以分为 5 大类。利用 RGA950作为探针进行Southern杂交，结果表明，其在基因组中存在多拷贝。【结论】本研究成功获得了不结球白 菜RGA序列，为进一步克隆不结球白菜的R基因奠定了基础。 关键词：不结球白菜；抗病基因同源序列；TIR-NBS-LRR；Southern杂交 Isolation and Characterization of Resistant Gene Analogs from Brassica campestris ssp.chinensis 1,2 1 2 2 2 WANG Yan-hua , HOU Xi-lin , SHEN Shu-xing , CHEN Xue-ping , WANG Mei 1 ( National Key Laboratory of Crop Genetics and Germplasm Enhancement, Nanjing Agricultural University, Nanjing 210095; 2 College of Horticulture, Hebei Agricultural University, Baoding 071001) Abstract: 【Objective 】Resistance gene analogs from Brassica campestris ssp.chinensis were isolated using homology-based method. 【Method 】The degenerated primers were designed according to the conservative domain TIR-NBS-LRR of most disease resistant genes in plants,and the genomic DNA and cDNA of B. campestris ssp.chinensis were amplified. 【Result 】Ten resistant gene analogs were obtained which contain the TIR-NBS-LRR domain. The deduced amino acid sequences have different homology scores compared with those of resistance genes or resistance gene analogs from other plants in