The mutant form of acetolactate synthase genomic DNA from rice is an efficient selectable marker for genetic transformation
The mutant form of acetolactate synthase genomic DNA from rice is an efficient selectable marker for genetic transformation
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DOI：
10.1007/s11032-005-0999-y
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发表时间：
2005-11-01
影响因子：
3.1
通讯作者：
Toki, S
中科院分区：
文献类型：
--
作者：
Osakabe, K;Endo, M;Toki, S
在转化过程中标记基因的正确使用对于转基因植物的生产至关重要。然而，消费者的关注和监管要求提出了反对在转基因植物中存在外源DNA，特别是来自病毒的抗病基因和启动子。克服这一问题的一种方法是通过使用几种位点特异性重组系统从植物基因组中消除标记基因。我们提出了一种替代方法来解决这个问题，使用标记基因完全来自宿主植物DNA。本研究克隆了水稻乙酰乳酸合成酶（ALS）基因的调控序列和编码序列，并对ALS基因进行了抗除草剂突变。将该构建体转移到水稻基因组后，用除草剂双草醚钠盐有效地选择转基因植物，该除草剂双草醚钠盐抑制野生型ALS的活性。我们还分析了水稻ALS基因启动子与gusA报告基因的调控特征，发现GUS在水稻幼苗地上部分和根尖中组成型表达。该标记系统仅由宿主植物DNA组成，并能够在单子叶作物植物水稻中进行有效选择。该筛选系统可用于其他作物品种的转基因植株的筛选，并有望获得公众认可。
The proper use of a marker gene in a transformation process is critical for the production of transgenic plants. However, consumer concerns and regulatory requirements raise an objection to the presence of exogenous DNA in transgenic plants, especially antibiotic-resistant genes and promoters derived from viruses. One approach to overcome this problem is the elimination of marker genes from the plant genome by using several site-specific recombination systems. We propose an alternative method to solve this problem using a marker gene exclusively derived from the host plant DNA. We cloned a genomic DNA fragment containing regulatory and coding sequences of acetolactate synthase (ALS) gene from rice, and mutagenized the ALS gene into a herbicide-resistant form. After transfer of this construct to the rice genome, transgenic plants were efficiently selected with a herbicide, bispyribac-sodium salt, which inhibits the activity of wild type ALS. We also analyzed the regulatory feature of the rice ALS gene promoter with the gusA reporter gene and revealed that GUS expression was observed constitutively in aerial parts of rice seedlings and root tips. The marker system consisted exclusively of host plant DNA and enabled efficient selection in a monocot crop plant, rice. The selection system can potentially be applied to generate transgenic plants of other crop species and can be expected to be publicly acceptable.