威宁短柱油茶MKK3基因克隆及生物信息学分析

Cloning and Bioinformatics Analysis of MKK3 Gene from Camellia saluenensis

【目的】分离克隆威宁短柱油茶MAPKK基因家族相关基因,为挖掘威宁短柱油茶抗逆相关基因奠定基础。【方法】利用RT-PCR扩增MAPKK基因全长cDNA序列,利用生物信息学分析软件对其开放阅读框、基本理化性质、磷酸化位点及蛋白互作等进行分析。【结果】克隆获得威宁短柱油茶CsMKK3基因,全长2 219 bp,包含1个1 557 bp的开放阅读框,编码518个氨基酸,蛋白的相对分子量为57 524.61 Da,理论等电点5.44,是无跨膜结构和信号肽的不稳定蛋白,定位于细胞核,含有多个磷酸化位点,与MAPK蛋白激酶家族的MPK2、MPK14、MPK1、MPK8、MPK5、MPK7及MPK6等蛋白存在相互作用;系统进化分析其与同科植物茶亲缘关系最近。【结论】克隆分离的CsMKK3基因符合MAPKK基因家族特征,属于该基因家族成员。

【Objective】In order to provide basis for exploring the genes related to stress tolerance of Camellia saluenensis,the relative genes from MAPKK family gene are isolated and cloned.【Method】The full-length cDNA sequence of MAPKK gene is amplified by RT-PCR,and its open reading frame,basic physical and chemical properties,phosphorylation sites and protein interaction are analyzed by bioinformatics analysis software.【Result】The full length of CsMKK3 gene by cloned was 2 219 bp,containing an open-reading frame of 1 557 bp,encoding 518 amino acid,the molecular weight of 57 524.61 Da and the theoretical isoelectric point of 5.44.The amino acid sequence prediction showed that it was an unstable protein without transmembrane structure and signal peptide,located in nucleus,and contained multiple phosphorylation sites.CsMKK3 protein interacts with MPK2,MPK14,MPK1,MPK8,MPK5,MPK7,MPK6 from MAPK protein kinase family.Phylogenetic analysis showed that the sequence was closely related to C.sinensis.【Conclusion】The isolated and cloned CsMKK3 gene meet the characteristics of MAPKK gene family,indicating that it is the member of MAPKK gene family.