桂花OfPSY、OfPDS和OfHYB基因启动子克隆及表达特性分析

【目的】探究高温和外源脱落酸对桂花Osmanthus fragrans类胡萝卜素生物合成的3个相关基因，包括八氢番茄红素合成酶基因(PSY)、八氢番茄红素脱氢酶基因(PDS)、β-胡萝卜素羟化酶基因(HYB)的调控作用，为阐释桂花类胡萝卜素代谢调控的机制提供研究基础。【方法】根据桂花基因组数据库的序列，从桂花品种‘堰虹桂’‘Yanhong Gui’中克隆OfPSY、OfPDS、OfHYB基因的启动子序列，并进行生物信息学分析，再构建PCAMBIA3301-LUC载体在烟草Nicotiana benthamiana中瞬时表达，结合高温(37℃)和200 mg·L^-1脱落酸处理，分析启动子活性。【结果】获得OfPSY、OfPDS、OfHYB基因的部分启动子，其长度分别为1 908、1 521及1 830 bp。作用元件分析表明：3个启动子中均存在TATA-box和CAAT-box等启动子基本元件、光响应元件、脱落酸响应元件以及MYB和MYC结合位点。此外，在OfPSY启动子中，存在赤霉素响应元件；在OfPDS启动子中，存在茉莉酸甲酯响应元件、赤霉素响应元件、厌氧诱导型...

Cloning and expression characterization of OfPSY, OfPDS and OfHYB gene promoters in Osmanthus fragrans

  Objective  This paper aims to investigate the regulation of high temperature and abscisic acid (ABA) on promoters of three carotenoid biosynthesis genes in Osmanthus fragrans including PSY, PDS, HYB, providing the research foundation for regulation of carotenoid biosynthesis in O. fragrans.   Method  According to the sequences in genome database of O. fragrans, promoters of OfPSY, OfPDS, OfHYB were cloned from an O. fragrans cultivar ‘Yanhong Gui’, which were used for bioinformatics analysis. Then, these promoters were inserted into PCAMBIA3301-LUC vectors and used for transient expression in tobacco(Nicotiana benthamiana) leaves under treatments of high temperature (37 ℃) and 200 mg·L?1ABA.   Result  Our research cloned the promoter of OfPSY, OfPDS and OfHYB from genomic database of O. fragrans in length as 1908 bp, 1521 bp and 1830 bp respectively. All three promoters contain promoter basic elements TATA-box and CAAT-box, light response element, ABA response element as well as the binding site of MYB and MYC. Moreover, there was gibberellins (GA) response element in OfPSYP; OfPDSP contains methyl jasmonate (MeJA) and GA response element, pathogen induction element as well as element involved in defense and stress responsiveness; OfHYBP contains hormone response elements as auxin, ethylene and MeJA, as well as low temperature response element and pathogen induction element. The assays of in transient expression in tobacco leaves showed that relative high temperature can activate the promoter activities of OfPSY, OfPDS and OfHYB, and ABA can activate the promoter activities of OfPDS and OfHYB.   Conclusion  High temperature and ABA probably affect the carotenoid accumulation in O. fragrans by regulating the promoter activities of carotenoid biosynthesis genes. [Ch, 4 fig. 5 tab. 28 ref.]