Overview
We have developed a simple method for the rapid and reproducible isolation of DNA from single flies for amplification by the polymerase chain reaction (PCR) (Saiki et al, Science 239: 487), and direct sequencing by asymmetric PCR (Gyllensten and Erlich, Proc. Nat. Acad. Sci. 85: 7652). The simplicity of this procedure means that the problem of contamination with other amplified or cloned DNA is greatly reduced. Sufficient DNA is obtained from one fly for a minimum of 50 PCR analyses, and the DNA is stable for at least one month in the refrigerator. A simple modification of this technique allows the isolation of DNA suitable for use in inverse PCR (Ochman et al, Genetics 120: 621-623). These methods substantially reduce the time involved in DNA isolation, and among other uses, allows the PCR to be used to monitor the segregation of an allele for which there is no phenotype or transposition of an unmarked P element (Engels et al. Cell 62: 515-525).
Greg Gloor and William Engels
Details
Details of this protocol, Single Fly Genomic DNA Prep For PCR, are located on a web site other than Biocompare Protocols.