A simple preparation of mycoplasmal DNA template for PCR from biological samples using effective surfactants.

H Kobayashi, G Munthali, C Miyamoto, T Morozumi, K Mitani, N Ito, H Shiono, K Yamamoto

Index: J. Vet. Med. Sci. 58(5) , 477-9, (1996)

Full Text: HTML

Abstract

To prepare mycoplasmal DNA template for PCR from biological samples rapidly and easily, surfactants which can solubilize cell membranes effectively were investigated. 3-[3-Cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS) was considered an effective surfactant. This surfactant could solubilize mycoplasma cell membrane without suppressing the polymerase reaction. In addition, proteinase K treatment played an important role in preparing mycoplasmal DNA template from a simulated biological sample. It was therefore considered that a combination of proteinase K- and CHAPS- added lysis buffer would be more useful in preparing mycoplasmal DNA template. We could detect PCR products by using the lysis buffer with a simulated lung emulsion sample containing mycoplasma organisms at 10(4) CFU per gram.