The long-term objective of this research is to develop primers for use in high stringency polymerase chain reaction (PCR) detection of mycoplasmas associated with human acquired immunodeficiency syndrome (AIDS). The primers will have commercial utility in analyzing the source, nature, and role of mycoplasma associated with the development of AIDS. The technological approach we will utilize in developing these primers could be used to derive PCR primers for other pathogens associated with opportunistic infections. In the first and second aims of this project we will use two distinct, but complementary, technologies to develop mycoplasma-specific primers. In the first aim, we will utilize a variation of PCR, called intergeneic length polymorphism PCR (ILP-PCR), to obtain fingerprints of the t-RNA gene complex present in mycoplasma. In the second aim, we will initially use a PCR-based amplification system, coupled with restriction digestion, to obtain diagnostic restriction-length fragment polymorphisms (RFLPs), a procedure which has been called mapped restriction endonuclease site polymorphisms (MRSP). The diagnostic PCR bands obtained in both methods will be sequenced in order to design highly specific PCR primers. In phase II of this grant, we will use the specific primers on clinical samples from AIDS patients.
