目的 探究杯茎蛇菰Balanophora subcupularis中五环三萜酯生物合成途径的关键酶基因，为其生物合成机制及蛇菰属药用植物资源开发利用奠定了一定的研究思路和理论基础。方法 对杯茎蛇菰花柄、根茎和花序3个组织进行转录组测序与分析，通过PfamScan搜索三萜酯生物合成候选酶基因，并构建了BsubOSCs基因的酵母表达载体，将其导入酿酒酵母Saccharomyces cerevisiae羊毛甾醇合酶基因（erg7）缺陷突变株GIL77中以表征其功能。结果 从杯茎蛇菰转录组中挖掘到可能参与五环三萜酯生物合成的相关酶基因共87条，其中大多数基因在花序中表达量最高，说明花序可能是五环三萜酯的主要合成部位。2条BsubOSCs基因的酶功能表征发现BsubOSC2能够催化2,3-氧化鲨烯生成β-香树脂醇，并没有检测到BsubOSC1的催化产物。结论 通过转录组分析发现了杯茎蛇菰五环三萜酯的生物合成关键酶基因，其中BsubOSC2为β-香树脂醇合成酶基因，是蛇菰素A生物合成的关键酶基因之一。为进一步解析蛇菰素等三萜酯化合物的生物合成途径提供了基因资源，促进了杯茎蛇菰分子育种的研究。

Objective To explore the key enzyme genes of the pentacyclic triterpene esters biosynthesis pathway in Balanophora subcupularis, which lays a certain research idea and theoretical foundation for its biosynthesis mechanism and the development and utilization of medicinal plant resources in the genus Balanophora. Methods Transcriptome sequencing and analysis was performed on the three tissues of the flowerstalk, rhizome and umbel of B. subcupularis, then searching the candidate enzyme genes involved in triterpene ester biosynthesis by PfamScan, constructing the yeast expression vector of BsubOSCs gene which was introduced into the Saccharomyces cerevisiae lanosterol synthase gene（erg7）-deficient mutant strain GIL77 for functional characterization. Results A total of 87 candidate genes involved in pentacyclic triterpene esters biosynthesis were identified from the transcriptome of B. subcupularis, most of them were highly expressed in umbel, suggesting that umbel may be the main site of synthesis of pentacyclic triterpene esters. The enzyme function of two BsubOSC genes were characterized, which was found that BsubOSC2 could catalyze 2, 3-oxidosqualene to form β-amyrin, but no catalytic product of BsubOSC1 was detected. Conclusion Based on transcriptome analysis, the key enzyme genes involved in pentacyclic triterpene esters biosynthesis were found in B. subcupularis, and BsubOSC2 is a β-amyrin synthetase gene, and is one of the key enzyme genes for the biosynthesis of balanophorin A. This study provides gene resources for further analysis of the biosynthesis pathway of triterpene ester compounds such as balanophorins, and facilitate the research on molecular breeding of B. subcupularis.

R286.12

云南特色植物提取实验室自主研究项目基金(2022YKZY004)