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园艺学报 ›› 2019, Vol. 46 ›› Issue (2): 385-396.doi: 10.16420/j.issn.0513-353x.2018-0319

• 研究报告 • 上一篇    下一篇

茶树CsAIL的克隆及在不同休眠阶段的表达分析

张伟富1,刘 莹1,3,孙冷雪1,王 璐1,曾建明1,杨亚军1,王新超1,韦朝领2,*,郝心愿1,*   

  1. 1中国农业科学院茶叶研究所,国家茶树改良中心,农业部茶树生物学与资源利用重点实验室,杭州 310008;2安徽农业大学茶树生物学与资源利用国家重点实验室,合肥 230036;3西北农林科技大学园艺学院,陕西杨凌 712100
  • 出版日期:2019-02-25 发布日期:2019-02-25
  • 基金资助:
    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(SKLTOF20160103);国家自然科学基金项目(31600563)

Cloning of CsAIL in Tea Plant and Its Expression Analysis During Winter Dormancy Transition

ZHANG Weifu1,LIU Ying1,3,SUN Lengxue1,WANG Lu1,ZENG Jianming1,YANG Yajun1,WANG Xinchao1,WEI Chaoling2,*,and HAO Xinyuan1,*   

  1. 1Tea Research Institute,Chinese Academy of Agricultural Sciences,National Center for Tea Improvement,Key Laboratory of Tea Biology and Resources Utilization,Ministry of Agriculture,Hangzhou 310008,China;2State Key Laboratory of Tea Plant Biology and Utilization,Anhui Agricultural University,Hefei 230036,China;3College of Horticulture,Northwest A & F University,Yangling,Shaanxi 712100,China
  • Online:2019-02-25 Published:2019-02-25

摘要: 从茶树不同休眠状态腋芽转录组中筛选出一条差异表达基因,经全长克隆和同源性分析证实该基因为与多年生植物休眠的形成和解除密切相关的AINTEGUMEN-LIKE (AIL)的同源基因,命名为CsAIL。生物信息分析表明,该基因包含1个1 872 bp的开放阅读框,编码623个氨基酸。编码的蛋白质分子质量为69.2 kD,理论等电点为6.6,是一种非分泌性蛋白;该蛋白有两个保守的AP2结构域,是植物中特有的广泛参与生长发育调节的AP2亚家族成员。亚细胞定位预测CsAIL蛋白不存在于叶绿体或者线粒体中,可能定位于其他细胞器;经同源性分析,在‘云抗十号’和‘舒茶早’全基因水平上分别鉴定了11个和12个AIL同源基因。系统进化树及序列保守性分析显示,CsAIL与‘云抗十号’CSA001743.1和‘舒茶早’TEA027750.1的进化关系最近,与已知的拟南芥AIL蛋白处于不同分支,但具有典型的保守结构域和已知的重要氨基酸位点。启动子序列分析显示该基因可能受生理节律、光及多种激素等信号共同调控。CsAIL在茶树顶芽、腋芽和茎中的表达量较高,叶片中较低,花中几乎不表达。在茶树冬季类休眠和生理休眠阶段,该基因的表达量在叶和越冬芽中均逐渐下调并维持在较低水平;在生态休眠及萌动阶段显著上调。分析其可能的下游调控基因CsCYCD3.2和CsCYCD6.1的表达,二者与CsAIL的表达模式高度一致,与越冬芽的休眠状态存在高度关联。本研究表明,CsAIL可能是参与茶树休眠调控的关键基因,与CsCYCDs共同在茶树年休眠—生长循环中发挥作用。

关键词: 茶树, 休眠, 越冬芽, AIL基因, 表达分析

Abstract: In tea plant,on the basis of previous transcriptome study on the axillary buds at different dormancy states,a differentially expressed gene related to dormancy regulation was identified,cloned and sequenced. This gene was highly homologous to AINTEGUMEN-LIKE(AIL),and named as CsAIL. Bioinformatic analysis showed that the CsAIL gene contains an open reading frame(ORF)in 1 872 bp,encoding 623 amino acid residues. CsAIL is predicted as a secretory protein with a putative molecular mass of 69.2 kD and theoretical isoelectric point of 6.6. CsAIL contained two conserved AP2 domains and should be grouped into AP2 subfamily,a superfamily closely involved in plant growth and development. Subcellular localization showed that CsAIL protein possibly locates the other organelles except chloroplasts and mitochondria. Homology analysis indicated that 11 and 12 AIL homologous genes were identified at the level of‘Yunkang 10’and‘Shuchazao’tea plant genome,respectively. Analysis of phylogenetic tree and sequence conservatism showed that CsAIL had the closest evolutionary relationship with‘Yunkang 10’CSA001743.1 and‘Shuchazao’TEA027750.1,and had different branches with known Arabidopsis AIL protein,but had typical conserved domain and known important amino acid sites. Promoter sequence cloning and analysis showed that CsAIL gene may be co-regulated by physiological rhythm,light and multiple hormone signals. Comprehensive expression analyses indicated that CsAIL had high expression levels in tea apical buds,axillary buds and stems,but relative low in leaves and extremely in flowers. At the stage of paradormancy,the expression of CsAIL was gradually down-regulated. A pretty low transcript level was maintained during endodormancy period. On the contrary,rapid expression increase was detected during ecodormancy and bud flush. Furthermore,the expression patterns of CsCYCD3.2 and CsCYCD6.1,putative downstream targets of CsAIL,were highly consistent with CsAIL and a high coefficient of association was observed between them. These results suggest that CsAIL may be a pivotal gene in bud dormancy regulation by modulating CsCYCDs among the dormancy-growth cycles in tea plant.

Key words: tea plant, dormancy, overwintering bud, AIL gene, expression analysis

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