甜瓜S-腺苷甲硫氨酸脱羧酶基因的克隆及白粉病诱导表达分析

摘要

In order to investigate the effect of S-adenosylmethionine decarboxylase (SAMDC) gene in melon resistance to powdery mildew,according to the previous results of EST sequences from eDNA-AFLP library and the melon genome sequence data,the SAMDC gene was isolated from Chinese wild melon clone ‘Yuntian930'by RT-PCR and designated as CmSAMDC (GenBank Accession No.KF151861).The mORF (main open reading frame) was 1 095 bp encoding 364 amino acids with a molecular mass of 40 kDa.The predicted protein sequence showed high similarity with Cucumis sativus and Citrofortunella microcarpa.The SDS-PAGE showed that the expression protein was a fusion protein with the molecular weight of 40 kDa,which perfectly matched the mass calculated from the amino acid sequence.Quantitative real-time PCR analysis indicated that the expression level of CmSAMDC reached a maximum at 48 hpi (hours post inoculation) that over 7-fold to 0 hpi,and the expression of CmSAMDC was also detected in tendril,root,leaf and stem.These results indicate that CmSAMDC gene may play protective roles in melon resistance to powdery mildew infection.%为探讨S-腺苷甲硫氨酸脱羧酶(S-adenosylmethionine decarboxylase,SAMDC)基因在甜瓜抵御白粉病菌中的作用,根据已知EST序列和甜瓜基因组数据库,在甜瓜抗白粉病品种‘Yuntian930’中克隆获得该基因的全长编码序列,命名为CmSAMDC (GenBank登录号为KF151861).生物信息学分析表明,CmSAMDC的主开放阅读框(mORF)长1 095 bp,编码364个氨基酸,预测分子量为40 kDa.聚类分析表明,CmSAMDC预测蛋白与黄瓜和四季橘中该蛋白的同源关系最近,并与其他双子叶植物聚为一类.原核表达分析表明,CmSAMDC以融合蛋白形式表达,相对分子量约为40 kDa,与预测一致.实时定量表达分析表明,CmSAMDC基因受白粉病诱导表达,在接种后48 h表达量达到峰值,为接种前的7倍,并且在甜瓜的根、茎、叶、卷须中均有表达.结果提示该基因可能参与了甜瓜的抗白粉病反应.