摘要
使用ClustalW和DNAMAN3.0分析了本实验室克隆的15个黄瓜抗病基因类似序列(RGA)之间以及与烟草的N基因、亚麻的L6基因和拟南芥的RPS2基因之间的同源性,并对这些RGA进行了PCR和Southern验证与分析。结果表明:15个黄瓜RGA中,核苷酸序列同源性最高的是CsRGA2、CsR-GA4和CsRGA5,其次是CsRGA6、CsRGA7、CsRGA8和CsRGA9,CsRGA1和CsRGA3也存在较高的同源性;其余的RGA同源性较低。在氨基酸序列上也表现了相同的特征。与N、L6和RPS2等抗病基因的产物之间同源性最高46%,最低22%。根据核苷酸序列设计了15对特异引物,用PCR方法验证了所有黄瓜RGA的存在,其中CsRGA3在黄瓜品种间表现出多态性。利用特异引物PCR方法合成了15个黄瓜RGA的地高辛探针,与EcoRⅤ酶解的黄瓜‘215’、‘129’和‘L18’的基因组DNA进行Southern杂交,证实了各个RGA在基因组上的存在,同时发现,CsRGA9、CsRGA11～CsRGA15都是多拷贝,CsRGA3是单拷贝,其余为双拷贝。
The homology among fifteen RGAs of cucumber cloned in authors＇ laboratory and the homology between the RGAs and N gene from tobacco, and L6 gene from flax, and RPS2 gene from Arabidopsis was analyzed using Clustal W and DNAMAN 3.0 program. The fifteen RGAs were identified by PCR and Southern hybridization. The results showed that the highest homology of nucleotide sequence existed between CsRGA2, CsRGA4 and CsRGA5. Higher homology existed between CsRGA6, CsRGA7, CsRGA8 and CsRGA9, also existed between CsRGA1 and CsRGA3, Less homology existed in the other RGAs. The amino acid sequence homologies of the products translated from the RGAs were also similar to the above. The highest homology, 46% , and the lowest homology, 22% , between the RGAs and three novel disease resistance genes, N, L6 and RPS2, were exhibited. The RGAs were identified using PCR with fifteen pairs of special primers based on the nucleotide sequence of the RGAs. CsRGA3 showed polymorphism in cucumber varieties. The fifteen digoxin （DIG） labeling probe, which produced by performing special primer PCR, hybridized with genomic DNA of 215, 129 and L18, digested by EcoRV, using Southern blot method. The existence of the RGAs in cucumber genome was certified. CsRGA9, CsRGAll - CsRGA15 had multi copies, respectively, CsRGA3 had single copy, the others had two copies.
出处
《园艺学报》
CAS
CSCD
北大核心
2007年第2期355-360,共6页
Acta Horticulturae Sinica
基金
黑龙江省博士后项目(LSZH-04088)
黑龙江省自然科学基金资助项目(C200526)
