番木瓜干旱胁迫相关CpDHN基因的克隆与原核表达  

Molecular Cloning and Prokaryotic Expression of CpDHN,a Gene Associated with Drought Stress in Papaya(Carica papaya L.)

作　　者：郭静远 邹智[2] 孔华 郭运玲[2] 郭安平[2] GUO Jingyuan;ZOU Zhi;KONG Hua;GUO Yunling;GUO Anping(School of Life and Pharmaceutical Sciences,Hainan University,Haikou,Hainan 570228,China;Hainan Key Laboratory for Biosafety Monitoring and Molecular Breeding in Off-Season Reproduction Regions/Institute of Tropical Bioscience and Biotechnology,Chinese Academy of Tropical Agricultural Sciences,Haikou,Hainan 571101,China)

机构地区：[1]海南大学生命科学与药学院,海南海口570228 [2]海南省南繁生物安全与分子育种重点实验室/中国热带农业科学院热带生物技术研究所,海南海口571101

出　　处：《热带作物学报》2020年第12期2406-2412,共7页Chinese Journal of Tropical Crops

基　　金：海南省重点研发计划项目(No.ZDYF2017017);中国热带农业科学院基本科研业务费专项(No.1630052017011)。

摘　　要：本研究基于转录组获得的CpDHN转录本序列,以番木瓜‘台农二号’组培苗叶片为材料,采用RT-PCR技术克隆了该基因包含完整ORF在内的425 bp cDNA序列。序列分析表明,CpDHN预测编码93个氨基酸,其理论分子量为10.50 kDa、等电点为6.62、总平均疏水指数为-1.984、核定位。除保守的K片段外,蛋白还含有1个S片段,可归为KS型脱水素。在拟南芥中的10个脱水素中,CpDHN与AtLEA8的亲缘关系最近,相似性为53.3%。值得注意的是,虽然CpDHN的编码区不存在内含子,但其3’UTR含有1个与AtLEA8类似的内含子。表达谱分析显示,该基因在根和叶片中均受干旱胁迫诱导。此外,还构建了CpDHN的原核表达载体,SDS-PAGE分析显示,该蛋白在大肠杆菌中可高效表达。这些结果为下一步的功能鉴定奠定了坚实的基础。Based on de novo assembled transcript sequences,a 425-bp cDNA of CpDHN,which includes the complete coding sequence,was isolated from the leaf tissue of papaya(Carica papaya L.) cultivar Tainong No.2 by using RT-PCR Sequence analysis revealed CpDHN putatively encodes 93 amino acids,which was predicted to target the nuclear,harboring a theoretical molecular weight of 10.50 kDa,an isoelectric point value of 6.62,and a GRAVY value of-1.984.CpDHN belongs to the KS-type dehydrin,which includes one conserved K-segment as well as one S-segment.Among ten family members present in Arabidopsis,CpDHN exhibits the highest sequence similarity of 53.3% with AtLEA8.Similar to AtLEA8, CpDHN contains no intron in the coding region but harbor one in the 3’UTR.Transcriptional profiling revealed that this gene was significantly regulated by drought stress in two of three tissues examined in this study,i.e.,root and leaf.Moreover,the prokaryotic expression vector of CpDHN was also successfully constructed,and SDS-PAGE showed that the protein could accumulate at considerably high level in Escherichia coli.Results obtained from this study provide a basis for further functional analysis.

关 键 词：番木瓜 脱水素 基因克隆 表达分析 原核表达 

分 类 号：S59[农业科学—作物学] S813.3