植物学报2025，Vol.60Issue(6)：875-887,13.DOI:10.11983/CBB24150
万寿菊花冠高效瞬时转化体系的建立及TeCYC2c基因启动子活性分析
Establishment of an Efficient Transient Transformation System for Tagetes erecta Corollas and Analysis on the Promoter Activity of TeCYC2c Gene
摘要
Abstract
INTRODUCTION:Marigold(Tagetes erecta),an important ornamental and medicinal plant,has a unique capitulum characteristic of the Asteraceae family with distinct ray and disc florets.However,the lack of efficient genetic transforma-tion system has limited the research on the mechanism of floral organ development of marigold.Floral transient trans-formation system offers a rapid approach to study the function of genes expressed specifically in floral organs.This study aimed to establish an efficient transient transformation system for marigold corollas and to analyze the promoter activity of TeCYC2c,which highly expressed in corollas,thereby laying the technical foundation for the rapid verification of floral gene function. RATIONALE:A fusion expression vector,incorporating the CaMV35S promoter and the GUS reporter gene,was con-structed to facilitate the transient transformation in marigold corollas.The study delved into the effects of bacterial strain type(GV3101,LBA4404,EHA105),bacterial suspension concentration(OD600 values 0.5-2.0),infection duration(10-40 min),and co-culture time(1-4 d)on the transient transformation efficiency of the GUS gene.Based on this transient transformation system for the marigold corollas,the promoter activity of the TeCYC2c gene was investigated.A 1 735 bp upstream sequence of the TeCYC2c gene was cloned and four promoter deletion expression vectors,with the GUS gene as the reporter gene,were constructed based on the distribution and quantity of elements predicted by PlantCARE.Subsequently,these vectors were employed for transient transformation of marigold corollas to facilitate an in-depth analysis of promoter activity. RESULTS:The transient transformation efficiency in marigold corollas demonstrated that the GV3101 strain achieved the highest infection efficiency;the bacterial suspension concentration,quantified at an OD600 value of 1.0,yielded the most robust transformation efficiency;the infection time was observed to exert no substantial influence on transient transformation efficacy;moreover,a co-culture time of 24 hours was identified as the optimal condition for the process.The results of GUS staining and GUS activity assay revealed that the core region of the promoter was located at-650 to-1 bp.It was speculated that the light-responsive elements within this region positively up-regulated the expression of downstream genes,while the hormone-responsive and stress-responsive elements unique to pTeCYC2c(-1 735)and pTeCYC2c(-1 406)might have an inhibitory effect on promoter-driven downstream gene expression. CONCLUSION:This study established an efficient transient transformation system for marigold corollas,optimized through strain selection and parameter tuning.The identification of the TeCYC2c promoter core region(-650 to-1 bp)and its regulatory elements provides critical insights into the regulatory mechanism of the TeCYC2c gene.The transient transformation system and promoter analysis method lay a technical foundation for accelerating functional studies of floral development genes in marigold,with potential applications in the genetic improvement of ornamental plants.关键词
万寿菊/TeCYC2c/启动子/瞬时转化/GUS活性Key words
marigold(Tagetes erecta)/TeCYC2c/promoter/transient transformation/GUS activity引用本文复制引用
窦淋琳,朱钰,刘翠翠,臧运平,陶正国,包满珠,何燕红..万寿菊花冠高效瞬时转化体系的建立及TeCYC2c基因启动子活性分析[J].植物学报,2025,60(6):875-887,13.基金项目
国家自然科学基金(No.32172616) （No.32172616）
