| 抗冻蛋白基因AFP表达载体构建及转化紫花苜蓿初报 |
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| 引用本文： | 郝凤,刘晓静,毛娟,于铁峰,张德罡. 抗冻蛋白基因AFP表达载体构建及转化紫花苜蓿初报[J]. 草地学报, 2009, 17(6): 779-783. DOI: 10.11733/j.issn.1007-0435.2009.06.015 |
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| 作者姓名： | 郝凤 刘晓静 毛娟 于铁峰 张德罡 |
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| 作者单位： | 1. 甘肃农业大学, 草业学院, 兰州, 730070;2. 甘肃农业大学, 农学院, 兰州, 730070 |
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| 基金项目： | 甘肃省农牧厅生物技术专项 |
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| 摘 要： | 以胡萝卜基因组DNA为模板,用PCR方法扩增目的基因,连接到PGEM-T Easy Vector载体上,经XbalⅠ和SacⅠ双酶切,将目的片段连接到PBI121工程质粒上,通过农杆菌介导法转化和田苜蓿,为提高紫花苜蓿的抗冻性奠定基础.结果表明,成功克隆出抗冻蛋白基因AFP,并构建成AFP植物表达载体.获得了具有卡那霉素抗性的苜蓿愈伤组织,和田苜蓿愈伤组织的最佳Kan筛选浓度为60 mg/L.经PCR技术鉴定,AFP抗冻蛋白基因已整合到苜蓿基因组中.
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| 关 键 词： | 抗冻蛋白基因AFP 载体构建 遗传转化 紫花苜蓿 |
| 收稿时间： | 2009-04-09 |
Construction of Antifreeze Protein Gene AFP Expression Vector and Transformation into Alfalfa Callus |
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| HAO Feng,LIU Xiao-jing,MAO Juan,YU Tie-feng,ZHANG De-gang. Construction of Antifreeze Protein Gene AFP Expression Vector and Transformation into Alfalfa Callus[J]. Acta Agrestia Sinica, 2009, 17(6): 779-783. DOI: 10.11733/j.issn.1007-0435.2009.06.015 |
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| Authors: | HAO Feng LIU Xiao-jing MAO Juan YU Tie-feng ZHANG De-gang |
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| Affiliation: | 1. Pratacultural College, Gansu Agricultural University, Lanzhou, Gansu Province 730070, China;2. College of Agronomy, Gansu Agricultural University, Lanzhou, Gansu Province 730070, China |
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| Abstract: | In order to enhance the freezing tolerance of alfalfa, the carrot genomic DNA was taken as template, the target gene was amplified using PCR method and connected to the PGEM-T Easy Vector vector, the target fragment was connected to the engineering plasmid PBI121 after double digestion by Xbal Ⅰ and Sac Ⅰ, then alfalfa Hetian was transformed through Agrobacterium-mediated transformation. The results show that the antifreeze protein gene AFP was successfully cloned, AFP plant expression vector was constructed, and the kanamycin-resistant callus of alfalfa was obtained. The optimal kanamycin selection concentration of alfalfa Hetian callus was 60 mg·L~(-1). Based on the identification by PCR technology, AFP antifreeze protein gene was integrated into the alfalfa genome. |
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| Keywords: | Antifreeze protein gene AFP Vector construction Genetic transformation Alfalfa |
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