荸荠CCoAOMT基因的克隆与表达分析  

Cloning and Expression Analysis of CCoAOMT Gene in Chinese Water-Chestnut

作　　者：陈夏鑫 陈振林[1,2] 帅良 段振华 商飞飞[1] 宋慕波[1] Chen Xiaxin;Chen Zhenlin;Shuai Liang;Duan Zhenhua;Shang Feifei;Song Mubo(Institute of Food Science and Engineering Technology,Hezhou University,Hezhou,542899;School of Food Science and Technology,Dalian Polytechnic University,Dalian,116034)

机构地区：[1]贺州学院,食品科学与工程技术研究院,贺州542899 [2]大连工业大学食品学院,大连116034

出　　处：《分子植物育种》2020年第20期6685-6691,共7页Molecular Plant Breeding

基　　金：国家重点研发计划(2018YFD0901003);国家自然科学基金(31801607);广西自然科学基金项目(2017GXNSFAA198082);广西聘专家专项经费(厅发[2016]21号)共同资助。

摘　　要：咖啡酰辅酶-A-O-甲基转移酶(CCoAOMT)是植物木质素合成的关键酶之一。为探明CCoAOMT在荸荠木质素单体合成过程中的作用,本研究利用RACE技术在荸荠球茎中克隆CCoAOMT基因的全长序列,并对其在不同组织和鲜切荸荠贮藏过程中的表达进行分析。结果表明,克隆得到荸荠CCoAOMT基因cDNA全长1 302 bp,将此基因命名为CwCCoAOMT,其GenBank登录号为MG719240。该序列包含一个786 bp的最大开放阅读框,可编码261个氨基酸,其DNA中包含4个内含子和5个外显子。该蛋白的分子量为29.17kD,理论等点为5.21。CCoAOMT蛋白二级结构中α-螺旋占37.55%,无规则卷曲占35.63%,延伸链占19.16%,β-折叠结构占7.66%。序列比对和系统进化树分析表明,Cw CCoAOMT蛋白与其它植物CCoAOMT蛋白序列相似度较高,其中与慈竹的亲缘关系最近。通过各组织表达特性分析发现,CwCCoAOMT在荸荠皮中的表达最高,在荸荠肉中的表达最低。在鲜切荸荠贮藏过程中CwCCoAOMT的表达呈快速上升趋势,而水杨酸处理能显著抑制CwCCoAOMT表达上升。本实验结果可为控制鲜切荸荠黄化的研究提供理论依据。Caffeoyl-coA O-methyltransferase(CCoAOMT) is a key enzyme of lignin biosynthesis in plants. In order to investigate the effects of the CCoAOMT on lignin biosynthesis in Chinese water chestnut(CWC), the full-length cDNA of CCoAOMT gene was cloned with RACE technology, and the expression of CCoAOMT gene in different tissues and in fresh cut CWC during storage was analyzed. The results indicated that, 1 302 bp full-length of CCoAOMT nucleotide was obtained from CWC c-DNA, and it was named CwCCoAOMT(GeneBank accession NO.MG719240). This gene contained an open reading frame of 786 bp, which encoded a polypeptide of 261 amino acids residues, and the DNA of CCoAOMT gene contains 4 introns and 5 extrons. The molecular weight of the predicted CCoAOMT protein was 29.17 kD, its theoretical p I was 5.21. The secondary structure of CCoAOMT protein consists of α-helix(37.55%), random coil(35.63%), extended strand(19.16%) and β-turn(7.66%).Sequence alignment and phylogenetic analysis showed that CwCCoAOMT shared high similarity with CCoAOMT proteins from other species, and very closely related to CCoAOMT protein of Bambusa emeiensis. Real-time PCR results showed that expression level of CwCCoAOMT was highest in peel and lowest in pulp. The expression of CwCCoAOMT increased dramatically during storage of fresh-cut CWC, while salicylic acid treatment significantly inhibited the expression of CwCCoAOMT. The results can provide a theoretical basis for the study of controlling the yellowing of fresh-cut Eleocharis dulcis.

关 键 词：荸荠(Eleocharis dulcis) CCoAOMT基因 基因克隆 生物信息学分析 表达分析 

分 类 号：S645.3[农业科学—蔬菜学] Q943.2[农业科学—园艺学]