新疆无苞芥类钙调素蛋白基因OpCML13的克隆与分析  

Clone and Expression Analysis of Calmodulin-Like Protein Gene OpCML13 fromOlimarabidopsis pumila of Xinjiang

作　　者：赵云霞[1] 魏艳玲[1] 黄先忠[1] 

机构地区：[1]石河子大学生命科学学院,农业生物技术重点实验室,新疆石河子832003

出　　处：《西北植物学报》2014年第3期431-437,共7页Acta Botanica Boreali-Occidentalia Sinica

基　　金：国家自然科学基金(31060149);石河子大学高层次人才启动项目(RCZX200902);新世纪优秀人才支持计划(NCET-12-1072)

摘　　要：通过对无苞芥幼苗cDNA文库的随机克隆测序分析，获得1条与拟南芥编码类钙调素13（calmodulin-likel3，CMLl3）高度相似的EST（GenBank登录号为JZl52285）。利用RT-PCR技术从无苞芥cDNA中克隆了该基因，命名为OpCMLl3，该基因开放阅读框（ORF）为447bp，编码148个氨基酸。生物信息学分析表明，OpCMLl3蛋白的二级结构含有4个ca^2＋结合基序（EF-hands）结构，是一个亲水蛋白，无信号肽，无跨膜区域，为非跨膜蛋白；同源建模发现该蛋白包含8个α-螺旋结构和10个β-转角。系统进化树分析表明，OpCMLl3编码产物与拟南芥AtCMLl3和AtCMI。14进化关系较近，属于同一进化分支。半定量RTPCR结果显示，OpCMLl3基因在无苞芥的不同组织中都有表达，在根中表达量最高；高盐胁迫处理6～24h，OpCMLl3基因的表达明显增强，随后逐渐恢复正常表达水平；低温、干旱和ABA处理6h后基因表达明显上调，并且一直保持较高的表达水平。研究表明，OpCMLl3基因在无苞芥逆境胁迫中可能起着重要作用。An expressed sequence of tags （EST） which was highly similar to the calmodulin-likel3 （CML13） gene of Arabidopsis was obtained, through sequencing of clones randomly selected from previ- ously constructed cDNA library of Olirnarabidopsis pumila leaves. This gene was cloned from leaf cDNA of O. purnila by RT-PCR technique, which was designated as OpCML13. Bioinformatics analysis showed that the open reading frame （ORF） of OpCML13 gene contained 447 bp encoding a peptide of 148 amino acids residues. The structural analysis of OpCML13 showed that it included four EF-hands motifs,and it is a hydrophilic protein,and no signal peptide and no transmembrane domain were found in OpCML13 protein indicating it is not a transmembrane protein. The tertiary structural analysis of OpCML13 through Swiss- model showed that it contained eight a-helixes and 10-13-turns. Phylogenetic tree revealed that OpCML13 showed closer genetic kinship with AtCML13 and AtCML14 of A. thaliana indicating that they belonged to the same evolutionary branch. Expression analysis by RT-PCR indicated that OpCML13 displayed a much broader expression range in different tissues in O. purnila, with a maximum expression in root. Under stress, expression of OpCML13 was induced with different kinetics and magnitude. The expression of OpC- ML13 was up-regulated as early as 6～24 h of salt treatment before rapidly returning normal level. OpC-ML13 mRNA significantly increased after cold,drought and ABA treatment for 6 h and remained elevated for at least 6 h. Our research indicated that OpCML13 played an important role in adversity stress.

关 键 词：无苞芥 类钙调素蛋白13 基因表达 抗逆 

分 类 号：Q785[生物学—分子生物学] Q786