西兰花表皮特异硫蛋白基因的克隆与表达分析

表皮特异硫蛋白在硫代葡萄糖苷代谢调控中起着重要作用,包括催化异硫氰酸酯生成环腈类和腈类物质。本研究以西兰花为试验材料,在克隆表皮特异硫蛋白基因BoiEPS的基础上进行序列分析和表达分析,并初步明确其在野油菜黄单胞菌侵染下的功能。结果表明,BoiEPS基因全长1 270 bp,有1个长度为238 bp的内含子;编码区全长1 032 bp,编码343个氨基酸,编码蛋白具有2个Kelch结构域。系统发育分析结果表明,BoiEPS与来自芜菁、甘蓝型油菜的EPS聚为一组,但与其他十字花科植物的EPS处于不同分支。实时荧光定量PCR结果表明,BoiEPS的表达受野油菜黄单胞菌的诱导,在24 h的表达量最大;BoiEPS的过量表达可显著提高西兰花对黑腐病的抗性,病程相关蛋白基因BoPR1的表达量在3个过表达株系均明显增加。本研究结果为后续开展西兰花-黑腐病抗性机理研究奠定了理论基础。

Isolation and Expression Analysis of Broccoli Epithiospecifier Gene

Epithiospecifier plays an important role in the regulation of glucosinolate metabolism, and it catalyzes isothiocyanates into epithionitriles and nitriles. In this study, our aim is to isolate an epithiospecifier gene from broccoli and clarify its sequence characteristics as well as its expression pattern challenged by Xanthomonas campestris pv. campestris (Xcc). Results showed that the full genomic DNA of BoiEPS was 1 270 bp in length, harboring a 238 bp intron. The length of its complete coding sequence was 1 032 bp, encoding 343 amino acids, and the deduced protein contained two Kelch domains. Phylogenetic analysis indicated that BioEPS was grouped with EPSs from Brassica rapa and B. napus into the same clade, however, EPSs from other cruciferous plants were found to be clustered in different clades. Quantitative real-time PCR results revealed that the expression of BoiEPS was induced by Xcc, and the highest expression level was observed at 24 h. Over-expression of BoiEPS in broccoli significantly enhanced its resistance to Xcc, and the expression of pathogenesis-related gene BoPR1 increased significantly. Results of our present study provided an insight into exploring the mechanism involved in black rot disease resistance in broccoli.