目的 研究山茱萸Cornus officinalis CoHMGR基因编码蛋白质的亚细胞定位以及不同浓度的外源激素对CoHMGR基因表达和活性成分含量的影响。方法 农杆菌菌液侵染烟草叶片,激光共聚焦倒置显微镜观察CoHMGR蛋白的亚细胞定位。不同浓度的茉莉酸甲酯和乙烯利喷施山茱萸2年生幼苗,实时荧光定量PCR(quantitative real-time PCR,qRT-PCR)测定不同时期CoHMGR基因表达水平,高效液相色谱(HPLC)检测莫诺苷、马钱苷含量。结果 激光共聚焦显微镜下观察到CoHMGR蛋白定位于内质网中。qPCR和HPLC结果显示200 μmol/L茉莉酸甲酯和250 mg/L乙烯利对目的基因表达水平和活性成分含量都有正向诱导效应。结论 CoHMGR蛋白定位于内质网,外源喷施200 μmol/L茉莉酸甲酯和250 mg/L乙烯利能显著提高CoHMGR基因表达量和莫诺苷、马钱苷含量,为深入解析CoHMGR基因功能奠定基础。

Objective To study the subcellular localization of the protein encoded by theCoHMGR gene ofCornus officinalis,and the correlation of spraying different concentrations of exogenous hormones on the expression ofCoHMGR gene and the content of active ingredients.Methods Agrobacterium infects tobacco leaves,and the subcellular localization of CoHMGR protein was observed under a confocal inverted laser microscope.Using different concentrations of methyl jasmonate and ethephon to spray two-year-old seedlings ofC.officinalis,quantitative real-time PCR (qPCR) was used to determine the expression levels ofCoHMGR gene at different periods after treatment,high performance liquid chromatography (HPLC) was used to detect the content of morroniside and loganin.Results The CoHMGR protein localization in the endoplasmic reticulum was observed under the laser confocal microscope.The results of qPCR and HPLC showed that 200 μmol/L methyl jasmonate and 250 mg/L ethephon had a positive induction effect on the target gene expression level and active ingredient content.Conclusion The CoHMGR protein is located in the endoplasmic reticulum.The exogenous spraying of 200 μmol/L methyl jasmonate and 250 mg/L ethephon can significantly increase the expression ofCoHMGR and the content of monoside and loganin,laying a foundation for in-depth analysis of the function ofCoHMGR gene.

R286.12

国家自然科学基金资助项目(U1404829);中央本级重大增减支项目(2060302);河南省自然科学基金项目(202300410151);河南省科技攻关项目(202102110156);河南省中药材产业技术体系建设专项资金资助;河南省中药材产业科技特派员服务团资助