白菜型油菜MPK12基因克隆及表达分析  

Clone and characterization of MPK12 gene from Brassica campestris

作　　者：李萍[1] 聂亭亭[1] 张腾国[1] 郑晟[1] 王娟[1] 毛玉珊 

机构地区：[1]西北师范大学生命科学学院,甘肃兰州730070

出　　处：《西北农林科技大学学报（自然科学版）》2017年第5期26-33,40,共9页Journal of Northwest A&F University(Natural Science Edition)

基　　金：国家自然科学基金项目(31460099;31160089);甘肃省自然科学基金项目(1208RJZA268)

摘　　要：【目的】克隆白菜型油菜‘陇油6号’MPK12基因的全长cDNA序列,研究其组织表达特异性,分析MPK12基因在低温、盐、ABA和H_2O_2处理下的表达情况,以阐明MPK12基因在油菜中的生物学功能。【方法】利用RACE技术克隆MPK12基因cDNA全长,并对其全长基因进行生物信息学分析;构建系统发育树,研究其与相似序列的同源性;利用实时荧光定量PCR方法,分析MPK12基因的组织表达特异性以及在低温、盐、ABA和H_2O_2逆境胁迫下的表达情况。【结果】油菜MPK12基因cDNA全长1 395bp,包括5′-UTR 69bp,3′-UTR 207bp,开放阅读框1 119bp,编码372个氨基酸,预测蛋白质分子量42.6ku,理论等电点为7.9,二级结构主要包括α-螺旋和不规则卷曲。多序列比对和系统进化分析表明,油菜MPK12与拟南芥AtMPK12具有很高的同源性,为90.7%。实时荧光定量PCR结果显示,MPK12基因在油菜根、茎、叶、芽和种子中均有表达,没有组织特异性;同时,该基因的表达受低温、盐、ABA和H_2O_2胁迫诱导。【结论】克隆得到油菜MPK12基因,其在油菜适应逆境胁迫过程中发挥作用。[Objective] The study aimed to isolate MPK12 from Brassica campestris Longyou 6,ana- lyze its tissue expression specificity and expression under stresses of cold,salt,ABA and H2O2 ,and clarify its biological functions. [Method] The full-length cDNA of MPK12 from Brassica campestris Longyou 6 was isolated by RACE technique and analyzed by bioinformatics. Phylogenetic tree was constructed for studying homology with similar sequences. RT-PCR method was used to analyze the tissue expression specificity and the expression of MPK12 under cold, salt, ABA and H2O2 stresses. [Result] The full-length cDNA of MPK12 was 1 395 bp,containing a 5＇-UTR of 69 bp,a 3＇-UTR of 207 bp,and a 1 119 bp opening reading frame. It had 372 amino acids with the molecular weight of 42.6 ku and theoretical isoelectric point of 7.9. The main secondary structure of the protein was a helix with random coils. Comparison of amino acid sequences and phylogenetic analysis indicated that this gene had high identities with Arabidopsis thaliana （90.7%）. RT-PCR analysis revealed that MPK12 was expressed in roots,stems,leaves,buds and seeds with almost no tissue specificity. The transcript level of MPK12 was increased in response to cold, salt, H2O2 and ABA stress. [Conclusion] MPK12 gene was isolated from Brassica campestris ,and it played an important role during cold, satt,ABA and H2O2 stresses.

关 键 词：白菜型油菜 MPK12基因 分子克隆 表达分析 

分 类 号：Q946[生物学—植物学] Q785