香蕉<i>MaAQP1</i>启动子诱饵载体及干旱胁迫酵母单杂交cDNA文库的构建

许奕; 李羽佳; 魏卿; 王安邦; 王笑一; 宋顺; 李敬阳

doi:10.19303/j.issn.1008-0384.2020.10.005

香蕉MaAQP1启动子诱饵载体及干旱胁迫酵母单杂交cDNA文库的构建

Constructions of Banana MaAQP1 Bait Vector and Drought-resistance cDNA Library

摘要

摘要:
目的香蕉MaAQP1能够提高植物的耐旱性，研究香蕉MaAQP1的相关特性可为了解其干旱胁迫响应机制奠定基础。
方法通过克隆MaAQP1的启动子，将启动子构建到pHIS2诱饵质粒上，转化酵母菌构建诱饵表达载体，同时构建干旱胁迫的香蕉（Musa acuminat L. AAA group cv. Brazilian）cDNA文库。
结果克隆获得1 362 bp的启动子序列，通过分析其顺式作用元件，结果显示启动子序列中共有72个顺式作用元件，包括了TATA-box和CAAT-box核心元件，ABA响应元件、MYB元件、MYC元件、ERE元件、MeJA响应元件、光响应元件以及分生组织响应元件等；成功构建了MaAQP1诱饵载体和干旱胁迫条件下的cDNA文库，文库库容为1.25×10⁷ CFU，插入片段平均在1 200 bp左右。
结论本研究克隆获得了香蕉MaAQP1启动子并构建了干旱胁迫酵母单杂交cDNA文库，为下一步运用酵母单杂交筛选MaAQP1互作的转录因子、解析MaAQP1响应干旱胁迫的作用机制奠定了基础。

Abstract:
Objective To clone MaAQP1 from a drought stressed banana plant using a constructed bait vector, and establish a cDNA library of the transformed drought-resistance gene from single-hybrids yeast cells.
Method The promoter of MaAQP1 from a banana plant (Musa acuminat L. AAA group cv. Brazilian) under draught-stress was constructed on the pHIS2 plasmid as the bait to be transformed into yeast cells for the gene cloning. A cDNA library of MaAQP1 in the yeast was subsequently established for the study.
Result The MaAQP1 bait vector was successfully constructed. The sequence of the 1 362 bp promoter was cloned and analyzed to show 72 cis-acting TATA-box and CAAT-box core elements as well as elements of MYB, MYC, ERE, and MeJA as well as those of ABA, light, and meristem responses, etc. A drought-resistance cDNA library of 1.25×10⁷ CFU in capacity with an average insert size of about 1 200 bp was established.
Conclusion The results provided a basis for screening transcription factors of the cloned MaAQP1 in the yeast single-hybrids to decipher the mechanism of MaAQP1 in response to drought for improving the stress resistance of plants.

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