‘红阳’猕猴桃突变体果实花青苷合成相关结构基因的表达分析

Expression analysis of structural genes related to anthocyanin synthesis in a mutant of ‘Hongyang' kiwifruit

【目的】探讨红肉猕猴桃发育过程中果实花青苷合成途径中相关结构基因的功能。【方法】以‘红阳’突变体‘HY-09’同一果实的中果皮和内果皮为试材,采用pH示差法测定了果实发育过程中花青苷含量的动态变化,利用实时荧光定量PCR技术检测了花青苷合成相关结构基因的表达水平,并应用Permut Matrix软件对基因的差异表达进行了不同部位和发育时期的双向分层聚类分析。【结果】整个果实发育过程中中果皮始终未能检测到花青苷的存在,而内果皮在花后67 d即有花青苷的积累,之后随果实发育进程花青苷含量逐步升高,至盛花后122 d时达最高值,随后至采收时稍有下降。对花青苷合成相关的结构基因进行qPCR检测分析,发现CHS2、CHI、F3H2、F3’H1、DFR2、ANS4、F3GT1和F3GGT1在中果皮和内果皮中都有表达,但其表达量和变化规律截然不同。所有基因在中果皮中的表达除盛花后47 d表达量稍高之外,其他时期都很微弱。内果皮中的CHS2、CHI、F3’H1、DFR2、ANS4、F3GGT1在果实发育前期表达量均较高,之后下降,至果实成熟时表达量最低;而F3H2和F3GT1的表达量高峰值出现在盛花后67 d,在果实发育后期仍保持较高量的表达,这与内果皮中花青苷积累模式相吻合。双聚类分析发现F3H2和F3GT1聚为一类,并与其他基因明显分开,说明2者的表达模式相似。【结论】红肉猕猴桃花青苷主要在内果皮中积累,相应地结构基因表达水平内果皮高于中果皮。且F3H2和F3GT1两个结构基因可能在猕猴桃花青苷的合成过程中起着重要作用,很有可能是花青苷合成的关键基因。

[Objective]The aim of this study was to explore the role of related structural genes in the anthocyanin synthesis in red-fleshed kiwifruit. [Method]The outer and inner pericarp in a single fruit of ＇HY-09＇ kiwifruit, a mutant of ＇Hongyang＇ （A ctinidia chinensis Planch. var. rufopulpa C. F. Liang et A. R. Ferguson） with stable red coloration were selected as materials. During the fruit development, the an- thocyanin content was determined by pH differential method and the expression of anthocyanin-synthesis- related structural genes were respectively analysed by qPCR （quantitative PCR） technique. [Result]The results showed that the anthocyanin can not be detected in outer pericarp during the whole fruit develop- ment. However, the anthocyanin was first detected at 67 day after full bloom （DAFB） in inner pericarp and then the anthocyanin content significantly increased along with the fruit development and peaked at 122 DAFB, but then decreased before harvest. Q-PCR analysis of structural genes showed that the level of gene expression in outer pericarp was very low except at 47 day DAFB during the fruit development. In in- ner pericarp, the expression levels of CHS2, CHI, F3＇H1, DFR2, ANS4, F3GGT1 were higher in young fruit period, but then decreased until kiwifruit .harvested The expression level of F3H2 and F3GT1 was highest at 67 day DAFB and remained high during the latter furit development, Hierarchical analysis suggested F3H2 and F3GT1 clustered together and separated from other genes significantly. [Conclusion] The anthocyanin was accumulated only in inner pericarp and the anthocyanin -synthesis- related structural genes had higher expression level than in the outer pericarp respectively. We suggested that F3H2 and F3GT1 might play important roles in the anthocyanin synthesis process, and it was likely to be key genes of anthocyanin synthesis in kiwifruit.