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茶叶科学 ›› 2017, Vol. 37 ›› Issue (6): 565-574.

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茶树Copine家族基因CsBON3的克隆与表达分析

姚利娜, 郝心愿, 王璐, 李娜娜, 曾建明, 杨亚军*, 王新超*   

  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,浙江 杭州 310008
  • 收稿日期:2017-01-18 修回日期:2017-04-23 出版日期:2017-12-15 发布日期:2019-08-23
  • 通讯作者: *
  • 作者简介:姚利娜,女,硕士研究生,主要从事茶树种质资源和育种研究。
  • 基金资助:
    国家自然科学基金(31370690)、国家茶叶产业技术体系(CARS-19)资助

Cloning and Expression Analysis of CsBON3 from Copine Gene Family in Tea Plant (Camellia sinensis)

YAO Lina, HAO Xinyuan, WANG Lu, LI Nana, ZENG Jianming, YANG Yajun*, WANG Xinchao*   

  1. Tea Research Institute of Chinese Academy of Agricultural Sciences/National Center for Tea Improvement, Key Laboratory of Tea Biology and Resource Utilization, Ministry of Agriculture, Hangzhou 310008, China
  • Received:2017-01-18 Revised:2017-04-23 Online:2017-12-15 Published:2019-08-23

摘要: Copine蛋白是一类包含2个C2(N端)和1个vWA(C端)保守域的Ca2+依赖蛋白或磷脂结合蛋白,在胞内信号转导中发挥重要作用。基于序列相似性分析,从茶树转录组数据库中筛选出1条与Copine家族基因高度同源的EST序列。经测序验证该序列包含1β746βbp的完整ORF,编码581个氨基酸。同源比对显示该基因与拟南芥AtBON3序列相似度最高(65%),将其命名为CsBON3(GenBank登录号为KY435900)。生物信息学分析显示,CsBON3蛋白分子量为63.66βkD,理论等电点为5.48;具有Copine家族蛋白特有的保守结构域;属亲水性蛋白,无信号肽位点,非分泌性蛋白,无跨膜结构域。表达分析表明,CsBON3在茶树花和根系中表达量最高,茎干和成熟叶中表达量最低。低温(4℃)处理茶树1βd后,其表达被显著上调;在生长阶段,该基因表达量高于休眠阶段;同时在接种炭疽菌的茶树叶片中,该基因也被快速上调,表明该基因可能与茶树低温、生长发育及抗病相关。

关键词: 茶树, Copine家族, BON基因, 逆境胁迫响应, 休眠, 表达分析

Abstract: Copine protein, a kind of Ca2+-dependent protein or phospholipid-binding protein containing two C2 (N-terminal) and one vWA (C-terminal) conserved domain, plays an important role in intracellular signal transduction. Based on the sequence identity analysis, an EST sequence showing high homology to Copine family genes was selected from our previous tea plant transcriptome database. The sequence was verified to contain 1β746βbp complete ORF, encoding 581 amino acids. Homologous alignment showed that the gene had the highest similarity with Arabidopsis AtBON3 (65%) and was named as CsBON3 (GenBank accession number: KY435900). Bioinformatic analysis showed that the molecular weight of CsBON3 protein was 63.66βkD and the theoretical isoelectric point was 5.48. It possesses the conserved domain unique to Copine family protein, which is a hydrophilic protein, no signal peptide site, non-secretory protein and no transmembrane domain. Expression analysis showed that CsBON3 had the highest expression levels in tea flowers and roots, and the lowest in stems and mature leaves. It was significantly up-regulated in tea plants treated with low temperature (4℃) for one day, and its expression was suppressed at dormant stage and up-regulated at growth stage. It was also up-regulated rapidly in the leaves with inoculation of Colletotrichum fructicola. The results indicated that the CsBON3 plays important roles in tea plant growth in response to low temperature and disease resistance.

Key words: tea plant, Copine family, BON gene, stress response, dormancy, expression analysis

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