黄瓜钙依赖性蛋白激酶基因CsCDPK5的克隆及响应淹水胁迫的表达分析  

Cloning and Expression Analysis of Cucumis sativus Calcium-dependent Protein Kinase 5 Gene(CsCDPK5) Under Waterlogging Stress

作　　者：许学文[1] 季晶[1] 陆璐[1] 齐晓花[1] 陈学好[1] 

机构地区：[1]扬州大学园艺与植物保护学院,江苏扬州225009

出　　处：《园艺学报》2016年第4期704-714,共11页Acta Horticulturae Sinica

基　　金：国家自然科学基金项目(31372087)

摘　　要：通过对黄瓜耐涝品系Zaoer-N和不耐涝品系Pepino淹水处理72 h后发现,Zaoer-N下胚轴平均可生成24.6条不定根,而Pepino平均仅能生成1.5条不定根,两者差异显著。从上述两个品种中克隆了钙依赖性蛋白激酶基因CsCDPK5的全长,两者序列完全一致,全长均为1 887 bp,编码629个氨基酸,预测其编码蛋白质分子量为59.17 k D,理论等电点为5.65。亚细胞定位预测表明CsCDPK5定位于细胞质。Zaoer-N和Pepino启动子相似度为99%,有14个碱基位点的差异,两者均含有与激素信号及低氧胁迫响应相关的顺式作用元件,而诱导子激活元件AT-rich sequence和功能未知元件F-box为Zaoer-N所特有的启动元件。qRT-PCR结果表明:(1)CsCDPK5在黄瓜下胚轴中的表达量最高;(2)淹水条件下,Zaoer-N和Pepino下胚轴中CsCDPK5的相对表达量均呈现先上升后下略有降,之后又上升的趋势,且Zaoer-N中的表达量大于Pepino;(3)根和叶中CsCDPK5的相对表达量无明显规律性;(4)在使用乙烯竞争性抑制剂1-MCP预处理后,淹水条件下Zaoer-N下胚轴不定根形成被抑制,CsCDPK5的表达与非淹水对照亦无明显差异。综上所述,黄瓜CsCDPK5为淹水胁迫响应基因,可能参与黄瓜淹水后下胚轴不定根的形成过程。Significant difference in adventitous root number （ARNs）was found between waterlogging tolerant line Zaoer-N and waterlogging sensitive line Pepino after 72 hours of waterlogging treatment, the average number of ARNs generated in Zaoer-N was 24.6, but only 1.5 in Pepino. Cucumis sativus calcium-dependent protein kinase 5 （CsCDPK5） was cloned from the two lines, which shared a sequence identity of 100%, the length of CsCDPK5 was 1 887 bp, and encoded 629 amino acids. It was predicted that the molecular mass of this protein was 59.17 kD, and pI was 5.65. CsCDPK5 protein was predicted to be localized in cytoplasm. The two lines shared a sequence identity of 99% in promoter sequence, and include 14 single nucleotide polymorphisms. Both of the two lines contained c/s-acting regulatory elements essential for phytohormone and anaerobic induction, whereas AT-rich sequence and F-box were special in Zaoer-N. The results of quantitative real-time PCR showed that： （1） CsCDPK5 was predominantly expressed in hypocotyls;（2） CsCDPK5 transcripts were upregulated at the beginning followed a slight down regulated in both hypocotyls of Zaoer-N and Pepino during waterlogging treatment, but higher in the former than latter; （3） CsCDPK5 transcripts were irregular in roots and leaves of both lines; （4） compared with controls, no significant changes were observed in CsCDPK5 transcripts when pretreatment with 1-MCP. In conclusion, CsCDPK5 is a waterlogging responsive gene and significant difference in transcripts were observed in hypocotyls of Zaoer-N and Pepino during waterlogging, and might play an important role in waterlogging-induced adventitious root formation.

关 键 词：黄瓜 淹水胁迫 不定根 CDPK 

分 类 号：S642.2[农业科学—蔬菜学]