Gel electrophoretic restriction fragment length polymorphism analysis of DNA derived from individual nematodes, using the PhastSystem

The DNA sequences constituting the internal transcribed spacer region, loca ted between 18S and 26S rDNA genes within the rRNA operon, derived from sin gle nematodes of two genera (Steinernema and Heterorhabditis) were amplifie d by polymerase chain reaction (PCR) and subjected to digestion by four res triction enzymes. The digests were analyzed by restriction fragment length polymorphism (RFLP) gel electrophoresis on the PhastSystem, using 7.5%, 5%C (Bis) polyacrylamide. The downscaling from conventional agarose to PhastSys tem gels permitted the analysis to be done on individual nematodes, rather than on mixed samples with average properties. The analysis time was reduce d so as to allow for the electrophoretic separation on 200 samples/workday. The resulting patterns of DNA fragments differed from those obtained by ag arose gel electrophoresis under conventional conditions by an increased num ber of detected fragments. The PhastSystem gel analysis provides the basis for taxonomical revisions (Pamjav et al., Electrophoresis 1999, 20 1264-127 1.).