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首页> 外文期刊>Journal of Biotechnology >A point mutation in the Medicago sativa GSA gene provides a novel, efficient, selectable marker for plant genetic engineering
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A point mutation in the Medicago sativa GSA gene provides a novel, efficient, selectable marker for plant genetic engineering

机译：苜蓿GSA基因中的点突变为植物基因工程提供了一种新颖，有效，可选的标记

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Bacterial selectable marker genes (SMG) conferring antibiotic resistance are valuable tools in plant genetic engineering, but public concern and regulatory requirements have stimulated the development of alternative selection systems. We have previously demonstrated that a mutated Synechococcus elongatus HemL gene encoding glutamate 1-semialdehyde aminotransferase (GSA) is an efficient SMG in alfalfa. In fact, GSA is irreversibly inhibited by gabaculine (3-amino-2,3-dihydrobenzoic acid), but the mutated enzyme is gabaculine insensitive. With the aim to develop a plant derived SMG, we cloned and sequenced the Medicago sativa GSA cDNA and reproduced one of the two mutations associated with gabaculine resistance in Synechococcus, a transversion resulting in a methionine to isoleucine (M. I) substitution. This mutated gene was assessed as a SMG in tobacco and alfalfa Agrobacterium transformation, in comparison with the wild type gene. In tobacco, about 43% of the leaf explants produced green shoots, whereas in alfalfa 47% of the explants produced green embryos in the presence of 30 mu M gabaculine when the M -> I GSA was introduced. Escapes were absent in tobacco and only 6% in alfalfa. No effect on the plant phenotype was noticed. We propose this new SMG as a widely acceptable alternative to those currently used
机译：赋予抗生素抗性的细菌选择标记基因（SMG）是植物基因工程中的重要工具，但公众的关注和法规要求刺激了替代选择系统的发展。以前我们已经证明，编码谷氨酸1-半醛氨基转移酶（GSA）的突变的细长突触球菌HemL基因是苜蓿中有效的SMG。事实上，GSA被加巴奎林（3-氨基-2,3-二氢苯甲酸）不可逆地抑制，但突变的酶对加巴奎林不敏感。为了开发植物衍生的SMG，我们克隆并测序了苜蓿GSA cDNA，并在Synechococcus中再现了与加巴奎林抗性相关的两个突变之一，这种突变导致蛋氨酸向异亮氨酸（M.I）的转化。与野生型基因相比，该突变基因被评估为烟草和苜蓿农杆菌转化中的SMG。在烟草中，当引入M-> I GSA时，在30μM加巴豆茶碱的存在下，约43％的叶子外植体产生绿芽，而在苜蓿中，则有47％的外植体产生绿胚。烟草中没有逸出，而苜蓿中只有6％。没有观察到对植物表型的影响。我们建议将此新的SMG作为目前使用的SMG的广泛接受的替代方案

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