MUTATOR TRANSPORTABLE ELEMENTS OF MAIZE

转座子在个体的一生中产生遗传多样性， 以及在进化的时间;他们的活动涉及许多 人类遗传疾病。 玉米是高等真核生物的典范 用于分析转座子的发育调控。 增变基因 转座因子家族是高等真核生物中最活跃的家族之一; 突变体线包含几十个移动的Mu元件，其插入 突变频率提高了100倍以上 突变转座子 活动在宿主发育过程中受到严格调节：Mu元件 切除、插入和拷贝数扩增都限于 组织分化的终末有丝分裂。 的结果 突变体的激活取决于组织：在索马中，元件切除， 但在生殖细胞中，元件扩增并插入（没有切除，因此 非常低的German Reversion频率）。 调节突变体元件 最近被克隆和测序;它编码两个收敛转录 具有共享终止区的基因。 使用“master”元素， 一方面，我们的长期目标是了解发展 突变体活动的调节以及宿主和 元素编码的活动决定了Mu的不同行为 体细胞和生殖细胞中的元素。 我们目前的目标包括 描述Mu 9，假定的“主”突变体元素，追求 三个具体目标：使用瞬时测定来表征更多 完整的两个基因的启动子、终止子和剪接; 通过分析Mu 9是一种“自主元件”的分子证据， 瞬时测定中的切除和转基因测定中的切除和插入 植物;并确定Mu 9基因产物在植物中的功能， 转座和/或元件表达。 利用最近分离的 具有新表型的突变体系-早期体细胞切除，高 基因突变的频率，和急剧的拷贝数减少--我们将 使用RNase保护分析来确定Mu 9的表达 在标准和变异突变体中进行比较。 这些数据应 提供深入了解突变体活动的发育调节。 利用小说系列大幅减少的拷贝数， 将测试一个简化的基因标记和克隆方案。

Transposons produce genetic diversity during the life of individuals as well as over evolutionary time; their activities are implicated in many human genetic diseases. Maize is a model higher eukaryote ideally suited for analyzing the developmental regulation of transposons. The Mutator transposable element family is among the most active in higher eukaryotes; Mutator lines contain dozens of mobile Mu elements whose insertion elevates mutation frequency greater than 100-fold. Mutator transposon activities are stringently regulated during host development: Mu element excision, insertion and copy number amplification are all restricted to the terminal mitotic divisions of tissue differentiation. The outcome of Mutator activation depends on the tissue: in the soma, elements excise, but in germ cells elements amplify and insert (without excision, hence a very low germinal reversion frequency). The regulatory Mutator element was recently cloned and sequenced; it encodes two convergently transcribed genes with a shared termination region. With the "master" element in hand, our long-term objectives are to understand the developmental regulation of Mutator activities and how the interplay of host and element-encoded activities determines the differential behavior of Mu elements in somatic and germinal cells. Our current goals include characterizing Mu9, the presumptive "master" Mutator element, pursuing three specific aims: to use transient assays to characterize more completely the promoters, terminators and splicing of the two genes; to provide molecular proof that Mu9 is an "autonomous element" by assaying excision in transient assays and both excision and insertion in transgenic plants; and to determine the functions of the Mu9 gene products in transposition and/or element expression. Using a recently isolated Mutator line with novel phenotypes -- early somatic excision, high germinal reversion frequency, and drastic copy number reduction -- we will use RNase protection analyses to determine how the expression of Mu9 compares in standard and the variant Mutator lines. These data should provide insight into the developmental regulation of Mutator activity. Capitalizing on the drastic copy number reduction of the novel line, we will test a simplified gene tagging and cloning protocol.