木薯ETR1基因克隆及表达分析  

Cloning and expression analysis of MeETR1 gene in cassava

作　　者：陈志晟 颜彦[2] 赵思涵 田丽波[1] 商桑[3] 胡伟[2] CHEN Zhi-sheng;YAN Yan;ZHAO Si-han;TIAN Li-bo;SHANG Sang;HUWei(College of Horticulture,Hainan University,Haikou 570228,China;Institute of Tropical Bioscience and Biotechnology,Chinese Academy of Tropical Agricultural Science,Haikou 571101,China;School of Life and Pharmaceutical Sciences,Hainan University,Haikou 570228,China)

机构地区：[1]海南大学园艺学院,海口570228 [2]中国热带农业科学院热带生物技术研究所,海口571101 [3]海南大学生命科学与药学院,海口570228

出　　处：《南方农业学报》2020年第1期19-26,共8页Journal of Southern Agriculture

基　　金：国家自然科学基金项目(31801419);海南自然科学基金面上项目(317036);海南省研究生创新科研项目(Hys2018-34)~~

摘　　要：【目的】克隆木薯乙烯受体基因(MeETR1)并检测其在木薯采后生理性变质(PPD)过程中的表达情况,为研究ETR基因家族在木薯PPD过程中的功能作用提供参考依据。【方法】以木薯品种华南8号为材料,应用PCR扩增MeETR1基因编码区(CDS)序列,采用生物信息学分析方法对其编码蛋白的理化性质、亲/疏水性、保守结构域及二、三级结构等进行预测分析,通过亚细胞定位确定MeETR1蛋白在植物细胞中的表达分布,并利用实时荧光定量PCR检测MeETR1基因在木薯采后PPD过程的表达情况。【结果】克隆获得MeETR1基因CDS序列全长2220 bp,共编码739个氨基酸,蛋白分子量为82.88 kD,理论等电点(p I)为6.76;MeETR1基因编码蛋白属于疏水蛋白,含有ETR1家族4个典型模块,即GAF结构域、HisKA结构域、HATPasec结构域和REC结构域;其二级结构中α-螺旋占47.43%、延伸链占14.46%、无规则卷曲占38.11%。MeETR1氨基酸序列(XP021625986.1)与同属大戟科的蓖麻(Ricinus communis)ETR1氨基酸序列(XP002533252.1)相似性为92.42%,亲缘关系较近;MeETR1蛋白定位在细胞质中。与0 h(PPD前)相比,MeETR1基因的相对表达量在木薯采后PPD过程中(12、24、48和96 h)均呈显著下降趋势(P<0.05),即MeETR1基因表达受木薯采后PPD过程的抑制。【结论】MeETR1基因编码蛋白含有GAF、HisKA、HATPasec和REC等4个典型的结构域,与其他植物ETR1在生物学功能上具有一致性;MeETR1基因表达在木薯采后PPD过程中受抑制,可能在此过程中发挥负调控作用。【Objective】In this study,the cassava ethylene receptor gene named MeETR1 was cloned and its expression level during the post-harvest physiological deterioration(PPD)process of cassava was detected,which laid a foundation for studying the function of the ETR gene family in cassava PPD process.【Method】The coding region(CDS)of MeETR1 gene was amplified from cassava variety SC8 by PCR method,and the physicochemical properties,hydrophilicity/hydrophobicity,conserved domains,secondary and tertiary structures,and phylogenetic relationship were analyzed by bioinformatics method.The distribution of MeETR1 protein in plant cells was determined by subcellular localization,and the expression pattern of MeETR1 gene in PPD process was detected by qPCR.【Result】The CDS of MeETR1 gene was 2220 bp in length and encoded 739 amino acids.The protein molecular weight was 82.88 kD,and the theoretical isoelectric point(pI)was 6.76.MeETR1 gene encoded protein was hydrophobin and contained four typical modules of the ETR1 family,namely GAF domain,HisKA domain,HATPasec domain and REC domain.In the secondary structure,theα-helix accounted for 47.43%,the extended chain accounted for 14.46%,and the irregular coil accounted for 38.11%.The amino acid sequence of MeETR1(XP021625986.1)was 92.42%similar to the amino acid sequence of Ricinus communis ETR1(XP002533252.1),which had the close genetic relationship.Subcellular localization proved that MeETR1 was localized in the cytoplasm.The relative expression level of MeETR1 showed a significant declined trend during PPD process(12,24,48 and 96 h)(P<0.05)compared with 0 H(before PPD),indicating MeETR1 gene was inhibited by cassava PPD process.【Conclusion】The protein encoded by MeETR1 gene contains four typical domains including GAF,HisKA,HATPasec and REC,and its biological function is consistent with other plant ETR1.MeETR1 gene is inhibited during the cassava PPD process and might play a negative role in regulating PPD of cassava storage roots.

关 键 词：木薯 乙烯受体(ETR) MeETR1基因 生理性变质(PPD) 表达分析 

分 类 号：S533.01[农业科学—作物学]