白芨钙依赖蛋白激酶基因BsCDPK1克隆及组织表达分析  

Cloning and Tissue Expression Analysis of Calcium-dependent Protein Kinase Gene BsCDPK1 in Bletilla striata

作　　者：王双双 曾钢 窦全丽[1] 张柏林[1] 田琴 Wang Shuangshuang;Zeng Gang;Dou Quanli;Zhang Bolin;Tian Qin(College of Biological and Agricultural Science and Technology,Zunyi Normal University,Zunyi,563006)

机构地区：[1]遵义师范学院生物与农业科技学院,遵义563006

出　　处：《分子植物育种》2021年第11期3571-3578,共8页Molecular Plant Breeding

基　　金：贵州省教育厅青年科技人才成长项目(黔教合KY字[2016]259号);遵义市科技局项目(遵市科合计[2018]11号);遵义市创新人才团队培养项目(遵市科人才[2018]1号);遵义师范学院博士启动基金项目(遵师BS[2015]12号);遵义师范学院赤水河流域资源保护与开发研究院开放基金项目(遵师CSHKF[2019]06号)共同资助。

摘　　要：钙依赖蛋白激酶(CDPKs)在调节植物生长发育与逆境胁迫响应中发挥了关键作用,研究CDPK基因的序列信息与表达模式,为揭示蛋白结构、基因功能及参与的抗逆信号网络奠定基础。采用RT-PCR、RACE技术获得白芨BsCDPK1基因c DNA全长;利用生物信息学软件分析其编码蛋白的理化特性和结构域等特征,并对BsCDPK1进行氨基酸序列比对与系统进化树分析;采用实时荧光定量PCR技术对Bs CDPK1基因在白芨幼苗期的表达模式进行检测分析。结果表明,克隆获得的Bs CDPK1基因cDNA全长为1981 bp,编码496个氨基酸,其编码蛋白相对分子质量为55.997 kD,理论等电点5.38,脂肪指数为87.50,不稳定指数为43.54。BsCDPK1蛋白包含CDPKs家族典型的激酶结构域、自抑制功能域、ATP结合位点以及C-端的4个EF-hand手性结构,并且高度保守;BsCDPK1与铁皮石斛、蝴蝶兰的CDPK基因亲缘关系最近,聚为一支。qRT-PCR结果表明BsCDPK1在白芨幼苗的根、茎、叶中均有表达。白芨BsCDPK1基因的克隆与序列特征分析、幼苗组织表达模式分析为阐明BsCDPK1基因在白芨生长发育和逆境胁迫中的功能奠定实验基础。Calcium-dependent protein kinases(CDPKs)play vital roles in the regulation of plant growth,development,and the response to adversity stress.Studying the sequence information and expression pattern of CDPK gene could lay the foundation for the protein structure,gene functions and signal network of plant stress resistance.The full-length cDNA of BsCDPK1 gene was obtained by RT-PCR and RACE approaches in this study.The physicochemical properties and structural domains of BsCDPK1 protein were analyzed by bioinformatics tools.In addition,amino acid sequence alignment and phylogenetic analysis of BsCDPK1 were performed.The expression pattern of BsCDPK1 gene in the seedling stage of Bletilla striata was detected by real-time fluorescence quantitative PCR.The result showed that the full c DNA of the cloned BsCDPK1 was 1981 bp,encoding 496 amino acids.The molecular weight of encoded protein was 55.997 kD,isoelectric point 5.38,aliphatic index 87.50 and instability index 43.54.The BsCDPK1 protein included the typical kinase domain of CDPKs in other plants,autoinhibitory domain,ATP binding site and the four EF-hands were highly conserved,which was more homologous with CDPKs of Dendrobium candidum and Phalaenopsis equestris,clustered on the same evolutionary branch.The results of qRT-PCR indicated that BsCDPK1 gene was expressed in the roots,stems and leaves in Bletilla striata seedlings.The cloning and sequence characterizing and tissue expression pattern analysis of BsCDPK1 provided experimental basis for elucidating the function of BsCDPK1 gene in plant growth and development and adversity stress responses.

关 键 词：白芨(Bletilla striata) 钙依赖蛋白激酶 基因克隆 表达分析 

分 类 号：S567.239[农业科学—中草药栽培]