新疆雪莲水孔蛋白sikPIP1基因的克隆与功能分析

doi:10.13560/j.cnki.biotech.bull.1985.2015.09.013

摘要/Abstract

摘要： 利用PCR技术从已建立的新疆雪莲cDNA文库中克隆雪莲水孔蛋白基因sikPIP₁，构建植物表达载体pBI121-sikPIP₁，利用农杆菌介导法获得转基因烟草，PCR和RT-PCR检测证明该基因成功导入并得以转录，并对检测结果均为阳性的植株通过水分胁迫和温度胁迫进行抗旱性和抗寒性分析。结果显示：(1)克隆得到长为880 bp，具有水孔蛋白特性的sikPIP₁基因，完整ORF为840 bp。(2)水分胁迫中，断水7 d后转基因烟草生长表型明显优于野生型烟草，生理指标测定结果显示，转基因烟草的相对电导率和丙二醛含量均低于野生型烟草，相对含水量高于野生型烟草。(3)不同温度胁迫处理，转基因烟草表型显著优于野生型，特别是0℃以下低温胁迫，野生型烟草出现严重的萎蔫，而转基因烟草受伤害程度较轻；生理指标结果显示转基因烟草相对电导率、丙二醛含量低于野生型烟草。结果表明，转sikPIP₁基因提高了烟草抗旱能力和抗寒能力。

关键词: 新疆雪莲, 水孔蛋白, sikPIP₁基因, 烟草, 抗寒性, 抗旱性

Abstract: Saussurea involucrata aquaporin protein gene sikPIP₁ was cloned from established cDNA library by PCR technique. Then a plant expression vector of pBI121-sikPIP₁ was constructed and transgenic tobacco plants were obtained by Agrobacterium-mediated method. The analysis by PCR and RT-PCR indicated that the aquaporin protein gene sikPIP₁ had been integrated and transcribed into the tobacco genome. The drought and cold resistance by water stress and low-temperature stress for the positive plants from above PCR and RT-PCR examination were analyzed. The results showed that：(1)The plasma membrane aquaporin gene sikPIP₁ was cloned with the length of 880 bp and complete ORF of 840 bp. (2)Under the water stress, the growth phenotype of transgenic tobacco was evidently better than wild-type after 7 days’ water-break, the results of physiological index showed that relative conductivity and MDA content of transgenic tobacco were lower than those of wild-type. However, the relative water content of transgenic tobacco was higher than that of wild-type. (3)The growth phenotype of transgenic tobacco was significantly better than that of wild-type under the different temperature stress. Especially below the 0 degree, the wild-type ones were wilting seriously, however, the transgenic tobacco had less damages. The results of physiological index showed that relative conductivity and MDA content of transgenic tobacco were lower than that of wild-type. The testing results showed that sikPIP₁ gene improved tobacco's tolerance of drought and cold.

Key words: Sasussured involucrate, aquaporin protein, sikPIP₁ gene, tobacco, drought tolerance, cold tolerance

黎玉顺, 刘逸泠, 刘步仓, 穆建强, 祝建波. 新疆雪莲水孔蛋白sikPIP₁基因的克隆与功能分析[J]. 生物技术通报, 2015, 31(9): 97-105.

Li Yushun, Liu Yiling, Liu Bucang, Mu Jianqiang, Zhu Jianbo. Cloning and Function Analysis of Aquaporin Protein Gene sikPIP₁ from Saussurea involucrata Kar. et Kir[J]. Biotechnology Bulletin, 2015, 31(9): 97-105.

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新疆雪莲水孔蛋白sikPIP₁基因的克隆与功能分析

Cloning and Function Analysis of Aquaporin Protein Gene sikPIP₁ from Saussurea involucrata Kar. et Kir

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