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作物学报 ›› 2008, Vol. 34 ›› Issue (03): 397-402.doi: 10.3724/SP.J.1006.2008.00397

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

昆虫抗冻蛋白基因转化烟草的抗寒性

王艳;邱立明;谢文娟;黄薇;叶锋;张富春;马纪*   

  1. 新疆生物资源基因工程重点实验室/新疆大学生命科学与技术学院,新疆乌鲁木齐830046
  • 收稿日期:2007-05-30 修回日期:1900-01-01 出版日期:2008-03-12 网络出版日期:2008-03-12
  • 通讯作者: 马纪

Cold Tolerance of Transgenic Tobacco Carrying Gene Encoding Insect Antifreeze Protein

WANG Yan,QIU Li-Ming,XIE Wen-Juan,HUANG Wei,YE Feng,ZHANG Fu-Chun,MA Ji*   

  1. Xinjiang Key Laboratory of Biological Resources and Genetic Engineering, College of Life Science and Technology, Xinjiang University, Urumqi 830046, Xinjiang, China
  • Received:2007-05-30 Revised:1900-01-01 Published:2008-03-12 Published online:2008-03-12
  • Contact: MA Ji

摘要: 将准噶尔小胸鳖甲抗冻蛋白基因MPAFP149亚克隆至pCAMBIA1302中,构建成单价植物表达载体pCAMBIA1302-MPAFP149,转化至农杆菌EHA105中,利用叶圆盘法转化烟草。通过PCR、PCR-Southern及RT-PCR检测表明抗冻蛋白基因已整合至烟草基因组中,并发生了转录。对T0代转基因烟草以-1℃处理48 h,转基因烟草的相对电导率和表型明显优于野生型烟草。室温恢复试验验证转基因烟草可存活并恢复生长,而野生型烟草受到了不可逆的低温冻害。研究证明转化后携带昆虫抗冻蛋白基因的烟草比野生型烟草具有明显的抗寒能力,该结果为减轻冷敏感经济作物在春季遭受霜害提供了理论依据和应用基础。

关键词: 昆虫抗冻蛋白, 植物表达载体, 烟草, FONT-FAMILY: 宋体, mso-ansi-language: EN-US, mso-fareast-language: ZH-CN

Abstract: Most of crops are susceptible to cold and frost. When temperature decreases to -1℃, ice makes plant cells desiccated and cellular membrane shrinked. Expression of insect antifreeze protein genes with very high thermal hysterisis activity in plants is a possible way for increasing the cold tolerance of cold-sensitive plants. The MPAFP149 gene from Xinjiang desert insect Microdera punctipennis dzhunarica was constructed to the plant expression vector pCAMBIA1302 by sub-cloning, and then the recombined vector pCAMBIA1302-MPAFP149 was transformed into Agrobacterium tumefacines EHA105. Transgenic tobacco was obtained via leaf-disc method by Agrobacterium-mediated transformation. PCR and PCR-Southern analysis showed that the MPAFP149 gene was successfully integrated into the tobacco genome. The result of RT-PCR also verified that MPAFP149 gene was transcripted at mRNA level. The relative conductivity of T0 generation of transgenic tobacco at -1℃ for 48 h was 28.83%, significantly lower than that of wild tobacco which was 82.91%. Meanwhile, the phenotype of transgenic tobacco was superior to that of wild tobacco, suggesting that transgenic tobacco had better cold tolerance than wild tobacco. Recovering experiment at the room temperature indicated that transgenic tobacco was able to recover from cold stress and regenerate. The results proved that transgenic tobacco carrying MPAFP149 gene was more cold tolerant than wild tobacco, which provided a theoretical and applied foundation for relieving cold damage to cold-sensitive crops in spring.

Key words: Antifreeze proteins, Plant expression vector, Tobacco, Cold tolerance

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