报春苣苔属植物花色变异的生化基础及转基因体系的构建

发布时间：2018-06-05 23:41

本文选题：报春苣苔 + 花色素苷　；参考：《仲恺农业工程学院》2017年硕士论文

【摘要】：报春苣苔属(Primulina)植物花色艳丽且变异丰富,整株花期较长,极具观赏性。本研究从遗传和发育两个角度分别对几种不同颜色的报春苣苔属植物及钟冠报春苣苔不同发育阶段的花色素苷成分和含量进行测定,揭示了报春苣苔花色艳丽的生化基础,并且以怀集报春苣苔为试材,建立了该属植物的转基因体系,为今后了解该属植物花色丰富变异的生理及遗传机制奠定了基础,有助于开展杂交育种及分子育种以进行花色种质创新。主要研究结果如下:(1)遗传方面,以38种报春苣苔属植物的新鲜花朵为试材,采用pH示差法测总花色素苷含量。结果显示花色越深,花色素苷的总含量越多,按照含量差异,从中选出13个物种用于花色素苷成分的鉴定,其中钟冠报春苣苔含有的花色素苷种类最多,为17种,白色的马坝报春苣苔和怀集报春苣苔及黄色的黄花牛耳朵不含有花色素苷,主要显色成分为黄酮类物质。(2)发育方面,以颜色最丰富的=钟冠报春苣苔为试材,通过花器官特异性研究发现,花瓣部分呈紫色是由于该部位蓝紫色的花色素(锦葵素、矮牵牛素及飞燕草素)的比例之和达到64.0%;而花筒部分的红色,是由于该部位矢车菊素及芍药花素分别占据了36.7%及20.0%。伴随着花发育过程,红粉色的花色素占据比例逐渐减小,而蓝紫色花色素所占比例渐趋增加,这可能是花瓣/花筒的比例伴随着花发育渐趋增大。开花前总花色素苷含量缓慢上升并趋于稳态(S2阶段有一峰值);而花盛开之后,含量急剧降低,这可能与花衰老进程的花色素苷降解相关。(3)转基因方面,以白色花的怀集报春苣苔为试材,利用农杆菌介导法建立了报春苣苔属植物高效快速转化体系,为花色育种提供条件。本实验以叶片为外植体进行植物组织培养,其中愈伤组织诱导培养基成分为MS培养基包含0.1 mg·L~(-1) TDZ和0.1mg·L~(-1) NAA,芽诱导培养基为MS培养基包含0.5 mg·L~(-1) TDZ和0.1 mg·L~(-1) NAA,所有的培养基均包含6 g·L~(-1)琼脂和30 g·L~(-1)蔗糖。恢复培养基中抗生素起始浓度为头孢噻肟500 mg·L~(-1),后期依次递减。用20 mg·L~(-1)的潮霉素经过4次筛选后进行PCR检测,转化效率为20%。
[Abstract]:The flowers of the genus Primulina were colorful and varied, and the whole plant had a long flowering period and was very ornamental. In this study, the anthocyanin composition and content of several different color plants of the genus Cesneria and Zhong Guan in different developmental stages were determined from the perspective of heredity and development respectively, and the biochemical basis of the brilliant color of the flower was revealed. Moreover, the transgenic system of this genus was established by using Cecilaria villosum as the material, which laid a foundation for understanding the physiological and genetic mechanism of the rich variation of flower color in this genus in the future. It is helpful to carry out cross breeding and molecular breeding to carry out flower color germplasm innovation. The main results were as follows: (1) in the aspect of heredity, the total anthocyanin content was measured by pH difference method with 38 fresh flowers of Cesneria. The results showed that the deeper the flower color, the more the total content of anthocyanin. According to the content difference, 13 species were selected for the identification of anthocyanin components, among which 17 species of anthocyanin were found in Zhong Guan. The ears of white and yellow cattle did not contain anthocyanin, and the main color component was flavonoids. The most colorful = Zhong Guan was used as the test material. Studies on flower organ specificity showed that the purple part of the petals was due to the sum of 64.0% of the anthocyanins (mallow, peachin and verdanin) in this part of the flower tube, while the red part of the flower tube was red. This is because cornwort and paeoniflorin occupy 36.7% and 20.0% respectively. With the development of flower, the proportion of red and pink anthocyanins decreased gradually, while the proportion of blue and purple anthocyanins gradually increased, which may be the proportion of petals / flower tube increased with the flower development. The content of total anthocyanin increased slowly before anthesis and tended to have a peak value in the stable phase of S 2, but decreased sharply after blooming, which may be related to the degradation of anthocyanin in the process of flower senescence. An efficient and rapid transformation system was established by means of Agrobacterium tumefaciens-mediated method. In this experiment, the leaves were used as explants for plant tissue culture. The callus induction medium consisted of MS medium containing 0.1 mg / L TDZ and 0.1mg L ~ (1) NAA, MS medium containing 0.5 mg / L ~ (-1) TDZ and 0.1 mg / L ~ (-1) NAA, and all medium containing 6 g / L ~ (-1) Agar and 30 g / L ~ (-1) sucrose. The initial concentration of antibiotics in the recovery medium was cefotaxime 500 mg / L ~ (-1), which decreased at the later stage. Hygromycin (20 mg / L) was used to detect PCR for 4 times, and the conversion efficiency was 20%.
【学位授予单位】：仲恺农业工程学院
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S68

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