SPECIFIC AMPLIFICATION OF SARCOCYSTIS-CRUZI DNA USING A RANDOMLY PRIMED POLYMERASE CHAIN-REACTION ASSAY

The polymerase chain reaction (PCR) method to randomly amplify polymor phic DNA (RAPD) was used to differentiate between Sarcocystis cruzi DN A and bovine DNA. This assay was also exploited to identify a S. cruzi DNA fragment which may be useful as a probe. Five primers ranging in length from 16 to 20 nucleotides were analyzed for their ability to di rect the amplification of either bovine or parasite DNA fragments. Two primers, TGA and TGD, preferentially amplified bovine DNA in a mixtur e of S. cruzi and bovine DNA. The primers TGB and TGF each directed th e amplification of S. cruzi DNA instead of bovine DNA. Assays using TG F and S. cruzi DNA resulted in the production of a unique 0.8 kilobase (kb) DNA fragment. This fragment was not amplified from two other clo sely related coccidian species, Toxoplasma gondii and Sarcocystis camp estris. When the 0.8 kb DNA fragment was purified and used as a DNA pr obe, it only hybridized with DNA from S. cruzi. The results of this st udy indicate that this DNA fragment may be developed into a useful DNA probe for S. cruzi, and that the RAPD-PCR method may be successfully exploited for the rapid development of DNA probes for parasites and ot her organisms.