2021年4月22日 星期四
茶树隐花色素基因CsCRY1和CsCRY2的克隆及表达模式分析
Cloning of cryptochrome gene CsCRY1 and CsCRY2 in Camellia sinensis and analysis on expression pattern
2020年 第29卷 第6期 页码[11-22]
摘要

通过同源比对茶树〔Camellia sinensis (Linn.) Kuntze〕基因组和转录组数据,获得2个茶树隐花色素(CRY)基因。利用PCR克隆技术,从茶树品种‘福鼎大白茶’(‘Fuding Dabaicha’)中克隆得到这2个隐花色素基因,分别命名为CsCRY1和CsCRY2。采用生物信息学方法,对茶树CsCRY1和CsCRY2基因编码蛋白的结构域、系统进化关系、二级结构和PHR结构域的三级结构进行分析,对CsCRY1和CsCRY2基因的启动子顺式作用元件及其功能进行了预测分析。结果显示:茶树CsCRY1基因开放阅读框长度为2 055 bp,编码684个氨基酸;CsCRY2基因开放阅读框长度为1 944 bp,编码647个氨基酸;CsCRY1和CsCRY2蛋白均在N端含有保守的PHR结构域,在C端含有保守的CCE结构域,这2个结构域在拟南芥〔Arabidopsis thaliana (Linn.) Heynh.〕中具有传导蓝光信号的功能。系统进化分析结果显示:CsCRY1和CsCRY2蛋白属于Plant CRY类隐花色素,其进化树分支距离与同目(杜鹃花目)植物滇山茶(Camellia reticulata Lindl.)和中华猕猴桃(Actinidia chinensis Planch.)较近,与单子叶植物距离较远。CsCRY1和CsCRY2基因的启动子顺式作用元件与光照和激素条件密切相关。实时荧光定量PCR分析结果显示:茶树根中CsCRY1和CsCRY2基因的相对表达水平最高,其后依次为叶、花、茎;蓝光能显著诱导CsCRY1和CsCRY2基因上调表达;0.1 mmol·L-1脱落酸(ABA)以及1.0 mmol·L-1吲哚乙酸(IAA)、茉莉酸甲酯(MeJA)和赤霉素(GA3)均能刺激CsCRY1和CsCRY2基因上调表达。研究结果显示:茶树CsCRY1和CsCRY2基因可能在组织发育、蓝光信号传导和激素调控中发挥重要作用。

Abstract

Two cryptochrome (CRY) genes in Camellia sinensis (Linn.) Kuntze were obtained by homologous comparison with genome and transcriptome datums of C. sinensis. These two cryptochrome genes, named CsCRY1 and CsCRY2, were cloned from C. sinensis ‘Fuding Dabaicha’ by using PCR cloning technology. Domain, phylogenetic relationship, secondary structure, and tertiary structure of PHR domain of proteins encoded by CsCRY1 and CsCRY2 genes in C. sinensis were analyzed by using bioinformatic method, and predictive analysis on cis-acting elements and their functions of promoters of CsCRY1 and CsCRY2 genes was carried. The results show that the open reading frame. of CsCRY1 gene in C. sinensis is 2 055 bp in length, which encodes 684 amino acids, and that of CsCRY2 gene is 1 944 bp in length, does 647 amino acids. There is a conserved PHR domain and a conserved CCE domain at N-terminal and C-terminal of CsCRY1 and CsCRY2 proteins, respectively, which have a function of transmitting blue light signals in Arabidopsis thaliana (Linn.) Heynh. The phylogenetic analysis result shows that CsCRY1 and CsCRY2 proteins belong to the cryptochrome of Plant CRY type, and their branch distances in phylogenetic tree are close to the same order (Ericales) plants of Camellia reticulata Lindl. and Actinidia chinensis Planch., but are far from monocots. The cis-acting elements of promoters of CsCRY1 and CsCRY2 genes are closely related to light and hormone conditions. The result of real-time fluorescence quantitative PCR analysis shows that relative expression levels of CsCRY1 and CsCRY2 genes are the highest in roots of C. sinensis, followed by leaves, flowers, and stems; blue light can significantly induce up-regulated expression of CsCRY1 and CsCRY2 genes; 0.1 mmol·L-1 abscisic acid (ABA), and 1.0 mmol·L-1 indole acetic acid (IAA), gibberellin (GA3), and methyl jasmonate (MeJA) can stimulate upregulated expression of CsCRY1 and CsCRY2 genes. It is suggested that CsCRY1 and CsCRY2 genes in C. sinensis may play important roles in tissue development, blue light signal transduction, and hormone regulation.

关键词茶树; 隐花色素; CsCRY1基因; CsCRY2基因; 表达模式
Key wordsCamellia sinensis (Linn.) Kuntze; cryptochrome; CsCRY1 gene; CsCRY2 gene; expression pattern
作者唐千惠a,b, 王佳欣a,b, 孙康a,b,c, 曾亮a,b,c, 吴致君a,b,c
所在单位西南大学: a. 食品科学学院, b. 食品科学与工程国家级实验教学示范中心, c. 茶叶研究所, 重庆 400715
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基金项目国家自然科学基金资助项目(31800586); 安徽农业大学茶树生物学与资源利用国家重点实验室开放基金项目(SKLTOF20190122)