菊花脑芳樟醇合酶基因CnTPS1的克隆与原核表达分析＊

目的 克隆菊花脑芳樟醇合酶基因CnTPS1的全长编码序列,利用原核系统表达融合蛋白,为进一步研究该基因在菊花脑萜类合成中的功能提供理论依据。方法 以菊花脑基因组数据为基础,设计特异性引物,PCR扩增CnTPS1的全长编码序列,利用生物信息学分析软件分析序列特征。利用qRT-PCR技术分析CnTPS1基因在不同组织中的基因表达量。构建原核表达载体,体外诱导目的蛋白表达。结果 CnTPS1编码序列全长1749bp,编码582个氨基酸;基因表达模式分析表明该基因在茎和管状花中表达量较高;原核表达系统能诱导出67.58kDa大小的蛋白。结论 首次从菊花脑中克隆得到一个芳樟醇合酶基因CnTPS1,运用生物信息学方法对其编码蛋白的理化性质、结构特征等进行了分析预测,分析了该基因的组织表达模式,并在原核表达系统中成功诱导表达出目标蛋白,这些结果将为菊花脑萜类合酶基因的功能以及萜类物质生物合成途径的解析提供理论依据。

Cloning and Prokaryotic Expression of the Linalool Synthase gene CnTPS1 from Chrysanthemum Nankingense

Objective To clone the full-length coding sequence of a linalool synthase gene CnTPS1 in Chrysanthemum nankingense, and express the fusion protein in a prokaryotic system to provide a theoretical basis for further study on the function of this gene in the synthesis of terpenoids in C. nankingense.Methods Based on the genome data of C. nankingense, specific primers were designed to amplify the coding sequence of CnTPS1. The sequence characteristics were analyzed by bioinformatics software. The expression levels of CnTPS1 gene in different tissues of C. nankingense were investigated by qRT-PCR. Prokaryotic expression vector was constructed and the expression of target protein was induced in vitro.Results The sequence was 1749bp in length, encoding 582 amino acids. The gene expression pattern analysis showed that the gene was highly expressed in stem and disc floret. Prokaryotic expression system can induce a protein with a size of 67.58kDa.Conclusion A linalool synthase gene CnTPS1 is cloned from C. nankingense for the first time. The physicochemical properties and structural characteristics of the protein encoded by CnTPS1 are analyzed and predicted by bioinformatics method. The tissue expression pattern of the gene is analyzed and the target protein was successfully induced in the prokaryotic expression system. These results will provide a theoretical basis for the analysis of the function of the C. nankingense terpene synthase gene and the analysis of the terpenoid biosynthesis pathway.