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

NSF植物基因组计划的目标是证明转座子ac(激活子)可以用作复杂玉米基因组中的基因鉴定和分离工具，以及诱变剂，并开发必要的工具来促进这一用途。第一个目标是在这个项目的初始阶段实现的，在这个阶段，ac被证明是专门插入或接近基因的，因此，在高度重复的玉米基因组中，ac是一个优秀的基因搜索引擎。培育了一个高度胚性的BZ WX自交系，并将其转化到Ac*和DS*结构中，并对其进行了修改，以便于转座子相邻序列的PCR分离。该项目第二阶段的主要目标是创建一套全面的转基因品系，作为未来插入文库生产的起点。这一阶段将分两个明确的阶段进行。第一个将展示工程的Ac*或Ds*元件的生发转座以及与转座子相邻的DNA的现成分离。第二个是确定一种玉米转化的方法，该方法将能够产生一组有用的转座子系，用于局部诱变。一旦实现了上述两个目标，将启动该项目的最后阶段，即在基因组中均匀分布的位置生产一组124个具有唯一标记元素的转基因株系。在这套方案中，玉米基因组中的大多数基因将位于发射平台的7厘米范围内，因此将成为局部转座子突变实验的现实目标。这些品系将被保存在玉米库存中心，并将作为感兴趣的科学家未来生成插入文库的开始材料。

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.