Fehon R G, Oren T, LaJeunesse D R, Melby T E, McCartney B M
Department of Zoology, Duke University, Durham, North Carolina 27708-1000, USA.
Genetics. 1997 May;146(1):245-52. doi: 10.1093/genetics/146.1.245.
Reverse genetic analysis in Drosophila has been greatly aided by a growing collection of lethal P transposable element insertions that provide molecular tags for the identification of essential genetic loci. However, because the screens performed to date primarily have generated autosomal P-element insertions, this collection has not been as useful for performing reverse genetic analysis of X-linked genes. We have designed a reverse genetic screen that takes advantage of the hemizygosity of the X chromosome in males together with a cosmid-based transgene that serves as an autosomally linked duplication of a small region of the X chromosome. The efficacy and efficiency of this method is demonstrated by the isolation of mutations in Drosophila homologues of two well-studied genes, the human Neurofibromatosis 2 tumor suppressor and the yeast CDC42 gene. The method we describe should be of general utility for the isolation of mutations in other X-linked genes, and should also provide an efficient method for the isolation of new allcles of existing X-linked or autosomal mutations in Drosophila.
果蝇中的反向遗传分析极大地受益于致死性P转座因子插入的不断积累，这些插入为鉴定必需基因位点提供了分子标签。然而，由于迄今为止进行的筛选主要产生常染色体P元件插入，该文库对于进行X连锁基因的反向遗传分析并不那么有用。我们设计了一种反向遗传筛选方法，利用雄性中X染色体的半合子状态以及基于黏粒的转基因，该转基因作为X染色体小区域的常染色体连锁重复。通过分离两个经过充分研究的基因（人类神经纤维瘤病2肿瘤抑制基因和酵母CDC42基因）的果蝇同源物中的突变，证明了该方法的有效性和效率。我们描述的方法对于分离其他X连锁基因中的突变应该具有普遍实用性，并且还应该为分离果蝇中现有X连锁或常染色体突变的新等位基因提供一种有效方法。
