Developmental Genetics Laboratory, London Research Institute, Cancer Research UK, 44 Lincoln's Inn Fields, London, UK.
Biotechniques. 2010 Jan;48(1):53-60. doi: 10.2144/000113348.
One of the many advantages of Drosophila melanogaster as a model organism is the relative ease with which gene deletions can be generated by imprecise excision of transposon insertions. Here, we describe a simple, fast, and efficient method of screening for single-gene excision events that is not biased by prior assumptions of the mutant phenotype. DNA sequence polymorphisms were used as co-dominant electrophoretic markers to identify candidate deletions in a single generation, and to delimit the breakpoints to within 0.5-1 kb, thereby rapidly identifying deficiencies that affect only the gene of interest. In addition, we used polymorphism profiling to map existing deficiencies. The method can also be applied to map the extent of deletions generated by x-rays and to identify targeted mutations generated by engineered zinc-finger nucleases in Drosophila and other polymorphic model organisms (e.g., zebrafish, mouse, Caenorhabditis elegans).
黑腹果蝇作为模式生物的众多优势之一，是通过转座子插入的不精确切除相对容易产生基因缺失。这里，我们描述了一种简单、快速、高效的筛选单基因缺失事件的方法，该方法不受突变表型的先验假设的影响。DNA 序列多态性被用作共显性电泳标记，以在单个世代中鉴定候选缺失，并将断点限制在 0.5-1kb 内，从而快速鉴定仅影响目标基因的缺陷。此外，我们还使用多态性分析来绘制现有缺陷。该方法还可用于绘制 X 射线产生的缺失程度，并鉴定由工程锌指核酸酶在果蝇和其他多态模式生物（如斑马鱼、小鼠、秀丽隐杆线虫）中产生的靶向突变。
