盐藻钙调素基因的克隆及表达分析  

Cloning and Expression Analysis of CaM from Green Alga Dunaliella salina

作　　者：王明芳 高相楠 徐微微 丛玉婷[1] 柴晓杰[1] WANG Mingfang;GAO Xiangnan;XU Weiwei;CONG Yuting;CHAI Xiaojie(Key Laboratory of Hydrobiology in Liaoning Province, Dalian Ocean University, Dalian 116023, China)

机构地区：[1]大连海洋大学,辽宁省省级高校水生生物学重点实验室,辽宁大连116023

出　　处：《水产科学》2021年第3期409-415,共7页Fisheries Science

基　　金：国家自然科学基金资助项目(31472260,30972240).

摘　　要：为探究盐藻钙调素基因的结构与功能,利用RT-PCR和RACE技术克隆到盐藻钙调素基因(DsCaM,GenBank登录号:MN428415),并对其进行生物信息学分析,通过qRT-PCR方法检测盐藻钙调素在盐胁迫条件下的表达情况。试验结果显示,盐藻钙调素基因cDNA全长1061 bp,开放阅读框495 bp,编码164个氨基酸。盐藻钙调素为亲水蛋白,该蛋白主要分布在细胞质和液泡内,无跨膜区域,不存在信号肽,蛋白质的二级结构以α-螺旋(56.71%)和无规则卷曲(26.22%)为主,成功构建蛋白质三级结构。系统进化分析表明,盐藻钙调素基因与莱茵衣藻钙调素基因亲缘关系最近。qRT-PCR结果表明,在高盐胁迫下盐藻钙调素基因的表达量显著上调,胁迫6 h时表达量达到最高值,差异达到极显著水平(P<0.01)。该研究成果将为进一步分析盐藻钙调素基因的功能及盐藻应答盐胁迫信号的分子途径提供新信息。In order to explore the structure and function of calmodulin gene in green alga Dunaliella salina,RT-PCR and RACE technology were used to clone D.salina calmodulin gene(DsCaM gene,GenBank accession number:MN428415).The DsCaM was then analyzed using bioinformatics method,and the expression of DsCaM gene under salt stress was detected by qRT-PCR method.The results showed that the full length of the cDNA of DsCaM gene was 1061 bp,the open reading frame was 495 bp,and it encoded 164 amino acids.DsCaM protein was hydrophilic and mainly distributed in cytoplasm and vacuoles,It contained no transmembrane area and signal peptide.The secondary structure of this protein was mainlyα-helix(56.71%)and random coils(26.22%).Further phylogenetic analysis showed that DsCaM gene and Chlamydomonas reinhardtii CaM genes were closely related.The results of qRT-PCR showed that the expression of DsCaM gene was significantly up-regulated under high salt stress,the maximum at 6 h(P<0.01).The findings take a significant role in further analysis of functions of the D.salina calmodulin gene and the molecular pathways under salt stress.

关 键 词：盐藻 钙调素 QRT-PCR 生物信息学分析 

分 类 号：S917.3[农业科学—水产科学]