目的 获得药用植物阳春砂Amomum villousm的萜类合酶基因AvTPS1的启动子并进行活性分析。方法 从阳春砂叶片基因组DNA（gDNA）中克隆获得AvTPS1基因，再通过FPNI-PCR的方法从阳春砂叶片gDNA中克隆AvTPS1的启动子并进行序列分析，构建由该启动子驱动β-葡萄糖苷酸酶基因（GUS）的重组表达载体pCAM-AvTPS1p，利用农杆菌介导法注射侵染烟草Nicotiana benthamiana叶片进行瞬时表达来验证AvTPS1启动子的活性。结果 获得AvTPS1的gDNA序列长度为2 444 bp，经过序列比对发现AvTPS1含有7个外显子和6个内含子；通过FPNI-PCR成功获得568 bp的AvTPS1启动子序列，其含有10种顺式作用元件，包括保守元件TATA-box和CAAT-box，以及可被转录因子MYC结合的G-box等；通过GUS染色发现AvTPS1启动子启动了GUS基因的表达，使转基因烟草叶片呈现蓝色。结论 AvTPS1启动子被成功克隆，且通过实验验证其具有启动基因表达的功能，为进一步研究AvTPS1在萜类合成途径的功能及其受转录因子调节的机制奠定了基础。

Objective To clone the unknown sequence of terpene synthase (TPS) AvTPS1 promoter from Amomum villousm and analyze its activity. Methods In this research, AvTPS1 DNA sequence was amplified and cloned from genomic DNA (gDNA) of A. villousm leaves. Furthermore, the promoter of AvTPS1 was cloned by FPNI-PCR and the sequence was analyzed. The recombinant vector pCAM-AvTPS1p with AvTPS1 promoter for the expression of GUS gene was constructed. The activity of AvTPS1 promoter was verified by transient expression of Agrobacterium-mediated infiltration using the leaves of Nicotiana benthamiana. Results The gene sequence of AvTPS1 was 2 444 bp including seven exons and six introns. The 568 bp AvTPS1 promoter was successfully cloned using FPNI-PCR. Furthermore the sequence had ten kinds of cis-elements including conserved elements TATA-box, CAAT-box, MYC2 related element G-box and other elements. Finally, the GUS staining showed the tobacco leaves infiltrated by the pCAM-AvTPS1p were blue. Conclusion The AvTPS1 promoter can drive the transcription of GUS gene and then it was verified to have the promoter activity. These results give foundation for future research on the function of AvTPS1 involved in the terpenoid biosynthesis and its relationship with the transcription factors.

R282.12

国家自然科学基金青年基金项目（81303163）；广东省高等学校优秀青年教师培养计划（Yq2013042）