目的 通过克隆鱼腥草乙酰辅酶A酰基转移酶(acetyl-CoA C-acetyltransferase,AACT)基因cDNA全长,并展开生物信息学分析和表达分析,为解析鱼腥草萜类次生代谢产物的生物合成机制奠定基础.方法 采用RT-PCR方法获得AACT基因cDNA序列并进行生物信息学分析;利用实时荧光定量PCR方法检测了AACT基因在鱼腥草的地下茎、地上茎、叶、花中的表达情况.结果 克隆获得的AACT基因cDNA全长为1 218 bp,编码405个氨基酸.生物信息学预测AACT蛋白不含跨膜区,不含信号肽.AACT基因在鱼腥草地上茎中的表达量最高,达到1.49,其次是地下茎0.96,花和叶中的表达量相对较低,分别为0.20和0.10.结论 首次从鱼腥草中克隆了AACT基因,为进一步阐明该基因在鱼腥草萜类化合物代谢途径中的重要作用奠定基础.

Objective Acetyl-CoA C-acetyltransferase (AACT) is the initial enzyme in the terpenoid biosynthesis pathway of mevalonate (MVA), two units of acetyl-CoA were catalyzed to acetoacetyl-CoA. To clone the full length cDNA of AACT gene and carry out the bioinformatics analysis and expression analysis in order to provide the basis on resolving the mechanism of biosynthesis for terpenoid secondary metabolites from Houttuynia cordata. Methods The cDNA sequence of AACT gene was obtained from H. cordata by using RT-PCR strategy. And the different expression of AACT gene in the rhizomes, stems, leaves, and flowers of H. cordata was analyzed by fluorescent quantitative PCR. Results The cDNA contains a 1 218 bp open reading frame and encodes a predicted protein of 405 amino acids. No transmembrane region and signal peptide were present in AACT protein by bioinformatics prediction. Relative real-time PCR analysis indicated that AACT gene showed the highest transcript abundance in the stems and rhizomes of H. cordata lower levels in the flowers and leaves, the values of them were 1.49, 0.96, 0.20, and 0.10, respectively. Conclusion This AACT gene is cloned from H. cordata for the first time. The results will provide a foundation for exploring the mechanism of the gene in terpenoid biosynthesis and metabolism in H. cordata.

国家自然科学基金资助项目(30870230);湖南省高校创新平台开放基金项目(12K132);湖南省科技计划重点项目(2013FJ6090)