A Set of Maize Transgenic Lines for Localized Mutagenesis Based on the Ac-Ds Transposon System Mutagenesis Based on the Ac-Ds Transposon System

这个NSF植物基因组项目的目标是证明转座子Ac（激活剂）可以用作基因鉴定和分离工具，以及诱变剂，在复杂的玉米基因组中，并开发必要的工具来促进这种使用。第一个目标在该项目的初始阶段实现，其中Ac被证明专门插入基因中或接近基因，因此是高度重复的玉米基因组中的优秀基因搜索引擎。开发了高度胚发生性bz wx近交系，并用Ac* 和Ds* 构建体转化，所述Ac * 和Ds* 构建体已被修饰以促进转座子邻近序列的PCR分离。该项目第二阶段的主要目标是建立一套全面的转基因品系，作为生产未来插入文库的起点。这一阶段将分两个明确界定的阶段进行。第一个将是证明工程化Ac* 或Ds* 元件的germinal转座和邻近转座子的DNA的容易分离。第二个将是确定一种玉米转化方法，该方法将能够产生一组有用的转座子系用于局部诱变。一旦达到上述两个目标，将开始项目的最后阶段，即生产一套124个转基因品系，这些品系在基因组中均匀分布的位置具有独特标记的元件。在该集合中，玉米基因组中的大多数基因将在发射平台的7cM内，因此将是局部转座子诱变实验中的现实靶标。这些株系将存放在玉米储备中心，并将作为感兴趣的科学家产生未来插入文库的起始材料。

The goals of this NSF Plant Genome project are to demonstrate that the transposon Ac (Activator) can be used as a gene identification and isolation tool, as well as a mutagen, in the complex maize genome, and to develop the necessary tools to facilitate that use. The first goal was met in the initial phase of this project where Ac was shown to insert exclusively in or close to genes and to be, therefore, an excellent gene-searching engine in the highly repetitive maize genome. A highly embryogenic bz wx inbred line was developed and transformed with Ac* and Ds* constructs that had been modified to facilitate the PCR isolation of the transposon-adjacent sequence. The main goal of the second phase of this project is to create a comprehensive set of transgenic lines that will serve as starting points for the production of future insertion libraries. This phase will occur in two well-defined stages. The first will be to demonstrate the germinal transposition of an engineered Ac* or Ds* element and the ready isolation of DNA adjacent to the transposon. The second will be to identify a method of maize transformation that will enable the production of a useful set of transposon lines for localized mutagenesis. Once the above two objectives have been met, the last stage of the project to produce a set of 124 transgenic lines with a uniquely marked element at evenly spaced locations in the genome will be initiated. In that set, most genes in the maize genome will be within 7 cM of a launching platform and will be, therefore, realistic targets in a localized transposon mutagenesis experiment. These lines will be deposited in the Maize Stock Center and will serve as starting materials for the generation of future insertion libraries by interested scientists.