一种能够抗多种除草剂的细胞色素P450基因的功能分析

Herbicide tolerant genes are the key component for the development of transgenic crops for herbicide tolerance. Due to the development of resistance to glyphosate by some weed species after long and overwhelming application, it is of great importance to develop novel transgenic crops that are tolerant to alternative herbicides. By rotating the application of glyphosate with other herbicides, it may greatly reduce the selection pressure for glyphosate tolerance on weeds. Recently, we have cloned a novel cytochrome P450 gene MR-1 from a weed species. This gene was found to be able to confer tolerance to a number of herbicides currently on markets in transgenic Arabidopsis. In this project we will determine the spectrum of herbicides that the transgenic plants with this gene confers to. MR-1 is highly similar to a corn cytochrome P450 gene that does not confer transgenic Arabidopiss to any herbicides tested. We will construct hybrid molecules by replacing parts of the MR-1 to its corn homologous gene with no herbicide tolerance activity. By evaluate the herbicide tolerance of these hybrid proteins, the project will likely be able to determine the motif in MR-1 which plays a key rule in determining the herbicide tolerance.

发掘抗除草剂基因是发展转基因抗除草剂农作物的关键。为了减缓杂草对草甘膦的抗性发展，目前迫切需要发展抗其他除草剂的新型转基因抗除草剂农作物。我们从一种杂草中克隆获得的一种能够抗多种除草剂的细胞色素P450基因MR-1。我们将详细分析和测定表达这种基因的转基因拟南芥对目前市场上常用除草剂的抗性能力，为进一步研发高效广谱抗除草剂转基因农作物提供基础抗性研究资料。同时本项目将通过构建MR-1与没有抗性的MR-1同源基因的杂交分子，分析和确定MR-1分子中决定抗除草剂性能的区域，为基因工程改良MR-1提供基础资料。

课题获得了一批转狗牙根P450-3X基因（申请书中命名为MR-1）的抗够除草剂的大豆。实验室和大田条件下试验表明，它们对如下除草剂具有明显的抗性能力：1）乙酰乳酸合成酶（ALS）抑制剂类除草剂：啶磺隆（Flazasulfuron）、烟磺隆（Nicosuflura）；2）对羟苯基丙酮酸盐双氧化酶（HPPD）抑制剂类除草剂：硝磺草酮(Mesotrione)；3）抗原卟啉原氧化酶（PPO）抑制剂类除草剂:氟磺胺草醚; 4）合成激素类除草剂：2,4-D和麦草畏(Dicamba)。因此，这些转基因大豆具有生产应用前景。此外，我们的研究发现了决定这种狗牙根P450蛋白质抗除草剂的决定性区域是一个65个氨基酸的多肽片段， 并且发现改变这个区域的氨基酸序列可以改变这个蛋白质的抗除草剂特性。