• 中文核心期刊
  • CSCD来源期刊
  • 中国科技核心期刊
  • CA、CABI、ZR收录期刊

CsZFP2基因的克隆及其在烟草中的抗旱功能分析

Connection between Tea ZFP2 and Drought Resistance of Tobacco Plants

  • 摘要:
      目的  对茶树ZFP2基因应答干旱胁迫表达进行分析,为进一步深入研究茶树ZFP2基因在植物生长发育与抵御干旱胁迫中的作用机制提供科学数据。
      方法  本实验室在前期工作中已通过酵母单杂筛选出一个与F3’5’H基因启动子区域互作的锌指蛋白,利用生物信息学的方法对茶树ZPF2的蛋白结构进行预测和分析,通过农杆菌介导法将CsZFP2基因转入烟草,在模拟干旱胁迫条件下进行功能验证。
      结果  茶树(Camellia sinensis,XM_028257969.1)锌指蛋白的蛋白结构域与烟草(Nicotiana tabacum,XP_016463943.1)的锌指蛋白相似性为68.57%。茶树锌指蛋白基因内含子数为13个,蛋白长度为1 233 a,蛋白分子量为141 204.45,等电点为5.9,等电点Pi<7表明茶树锌指蛋白富含酸性氨基酸;CsZPF2编码蛋白质主要二级结构为α螺旋且该蛋白是由α-螺旋和β-转角组成无规则卷曲的三维空间结构; CsZPF2编码蛋白质亚细胞定位在细胞核且氨基酸序列小于零部分多于50%,为亲水性蛋白;在自然干旱胁迫28 d后野生型烟草与转基因烟草底部老叶均明显发黄,但野生型烟草底部老叶基本完全失绿,呈枯萎状态,而转基因烟草底部叶片呈轻度萎蔫状态,大部分仍然呈现绿色。
      结论  CsZFP2基因能够响应干旱胁迫,并在干旱胁迫应答中起到作用,过表达CsZFP2的转基因植株比野生型植株更具抗旱能力。

     

    Abstract:
      Objective  Expression of tea ZFP2 in tobacco plants in response to drought stress was studied to elucidate the molecular mechanism.
      Method   A zinc finger protein (ZFP) was previously found in our laboratory to interact with the promoter region of F3'5'H gene. Using bioinformatics methods, the structure of ZPF2 from Camellia sinensis was predicted and analyzed. Then, CsZFP2 was cloned and transferred into tobacco by an Agrobacterium-mediated method for functional verification under a simulated drought stress.
      Result  The domain of the ZFP from C. sinensis XM_028257969.1 was 68.57% homologous to that from Nicotiana tabacum XP_016463943.1. There were 13 introns in CsZFP2 with a length of 1,233a, a molecular weight of 141 204.45, an isoelectric point of pH 5.9, and an isoelectric point Pi<7 indicating high content of acidic amino acids. The gene encoded mainly two proteins with an α-helix primary structure and a random coil 3D structure composed of α-helix and β-turn. The hydrophilic protein was subcellularly located in the nucleus with more than 50% of the amino acids having a sequence less than zero. Under natural drought for 28 d, the old leaves at the bottom of both wild-type and transgenic tobacco plants were turning yellow. However, those leaves on the wild-type plants became basically chlorotic and withered, but those on the transgenic plants wilted only slightly and largely remained green in color.
      Conclusion  CsZFP2 affected the response of tobacco plants to drought stress. The transgenic plants with overexpressed CsZFP2 were more resistant to the stress than the wild-type counterparts.

     

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