New Genetic system for the study of the plasid genome

我们将利用烟草，一个合适的模式物种，开发将基因导入高等植物叶绿体基因组的工具。高等植物中缺乏叶绿体转化的原因是细胞中的叶绿体基因组拷贝数太大，缺乏合适的选择标记，以及缺乏有效的DNA传递方式。首先，我们将用对抗生素产生抗药性的16S rRNA基因转化野生型叶绿体。DNA的引入将通过DNA包裹的钨颗粒的轰击来实现。转化将导致基因替换。研究将扩展到研究将外源DNA插入到叶绿体基因组中的要求。寡核苷酸和基因大小的DNA片段将与选择标记相连。寡核苷酸的掺入将被用来选择合适的基因插入位置。更大的DNA片段将包括嵌合基因。复制校正的参与将在稳定的转基因叶绿体基因组的形成中确定。从拟议的研究中获得的信息将使转基因植物中叶绿体基因调控和功能的研究成为可能。这项研究将为高等植物制定一种叶绿体转化方案。这是进一步进行植物基因工程的一个非常重要的工具。

We shall develop the tools for introducing genes into the plastid genome of higher plants using Nicotiana tabacum, a suitable model species. Lack of plastid transformation in higher plants has been due to problems of a large plastid genome copy number in a cell, lack of appropriate selective markers, and an effective way of DNA delivery. First, we shall transform wild-type plastids with plastid 16S rRNA genes that confer resistance to antibiotics. Introduction of DNA will be by bombardment with DNA-coated tungsten particles. Transformation will result in gene replacement. Research will be extended to study the requirements of inserting foreign DNA into the plastid genome. Oligonucleotide, and gene- size segments of DNA will be linked to the selective markers. Incorporation of oligonucleotides will be used to select suitable sites for gene insertion. Larger segments of DNA will include chimeric genes. Involvement of copy correction will be determined in the formation of stable transgenic plastid genomes. Information derived from proposed research will make feasible the study of plastid gene regulation and function in transgenic plants. This research will develop a plastid transformation protocol for higher plants. This is a very important tool for further genetic engineering of plants.