槜李等15个李品种S基因型鉴定及其多态性分析

利用李属S-RNase基因特异性引物,对15个供试李品种进行PCR扩增,共获得30个目的条带。对这些目的条带进行测序鉴定出15个李品种的S基因型。通过与NCBI中利用BLASTn与GenBank+EMBL+DDBJ+PDB等数据库中的序列比对,结果表明,其中9个为新S-RNases基因,对9个新S-RNases核苷酸序列进行分析发现,位于高变区内的内含子大小为141～1758bp,其同源性为33.9%(S-18～S-19)～81.6%(S-20～S-21),表现出丰富的长度和序列多态性;编码区的核苷酸序列比对结果,其同源性为73.3%(S-16～S-19)～91.7%(S-17～S-22);其推导氨基酸序列相似性为67.3%(S-16～S-19)～89.1%(S-17～S-22);包含李属S-RNase一级结构所共有的C2、C3保守区和高变区(RHV)。系统进化分析表明,9个新S-RNases与李属其它树种S-RNases聚类在一起,归属为李亚科(Prunoideae)。

Identification of S-genotypes of Japanese plum cultivars by PCR and analysis on polymorphism of S-RNase genes

PCR was conducted with Prunus S-RNase-gene-specific primers, which designed from the conserved regions of Rosaceous S-RNases and gave fragments comprising conserved regions C2 to C3, using 15 cultivars of Japanese plum. Each cultivar had two fragments. 30 fragments in all were cloned and sequenced. Analysis of these sequences indicated that 9 novel S-RNase genes were identified. The introns of 9 novel S-RNases located in the hypervariable region whose sizes ranged from 141 bp to 1 758 bp and the similarities exhibited from 33.9%(S-18—S-19) to 81.6% (S-20—S-21). The similar scores of the deduced DNA coding region and amino acid sequences of 9 novel S-RNases were 73.3% (S-16—S-19) to 91.7% (S-17 — S-22) and 67.3% (S-16—S-19) to 89.1% (S-17—S-22), respectively, carried the typical structure features of Prunus S-RNase: two conserved regions C2 and C3, one hypervariable region (RHV). Based on the DNA sequence analysis, the S-genotypes of 15 cultivars were determined as followed: Fugongqingpili (S-11/S-7),95-03 (S-27/S-a),Pingguoli(S-15/S-16), Kuandiandali (S-15/S-16), Yingtaoli (S-11/S-8),Xiguali(S-k/S-h), Zuili(S-h/S-7),Zhenzhuli (S-21/S-22),Zhuxueli (S-21/S-22), Qianping(S-10/S-i),Qiuli(S-e/S-11),Lihesihao (S-e/S-11),Meiguodali (S-11/S-20), Xinjiangsanhao (S-17/S-19),Tanchengxingmei (S-18/S-20). Phylogenetic analysis of 9 novel S-RNases and those of other Rosaceae species resulted in two distinct groups that correlate with their subfamily classification: Maloideae and Prunoideae, 9 novel S-RNases belong to the latter.