NATURE SCREEN - AN EFFICIENT METHOD FOR SCREENING NATURAL-POPULATIONSOF DROSOPHILA FOR TARGETED P-ELEMENT INSERTIONS

The efficiency of molecular techniques is making it increasingly neces sary to rely on reverse genetics to understand the function of genes. Tissue specific libraries allow one to identify numerous genes that ca n be cloned, sequenced, and mapped and whose temporal and tissue-speci fic pattern of expression are well characterized but whose function re mains unknown. In such cases, it is desirable to generate targeted mut ations to examine the phenotype of loss-of-function lesions. Here we d escribe a method for identifying naturally occurring variants of Droso phila melanogaster with specific genes tagged by a nearby P element. I mprecise P-element excision can then be used to generate a series of s mall deletions in or near the gene. In the method described here, larg e numbers of wild-caught males were crossed to balancer females, and i nserts were identified in pooled samples by the polymerase chain react ion with one primer from each target gene and one primer from the P-el ement terminal repeat. We present the calculations for the probability of successfully tagging a gene and show that it is greatly improved b y simultaneously screening inserts into several genes. If a large natu ral population is available, a nature screen is faster and easier than inducing P-element transposition in the laboratory, but the resulting lines, being genetically heterogeneous, may require more subsequent w ork to isolate. Using this method to screen the genomes of approximate to 10,400 males, we found P-element inserts in close proximity to 3 o f 10 genes that were screened.