Effect on Sensitivity of PCR Amplification of Ribosomal DNA Gene Using Universal Primer with a Mixture of Yeast Species

CHIN-CHENG HUANG^1*, WEI-CHUNG TSAI¹ AND HSI-HUA WANG²

1. Food Industry Research and Development Institute, Hsinchu, Taiwan, R.O.C.
2.Department of Agricultural Chemistry, National Taiwan University, Taipei, Taiwan, R.O.C.

ABSTRACT

   To detect the sensitivity of PCR using universal primers to amplify the RNA gene of internal transcribed spacer (ITS I) with a mixture of yeast species. The amounts of extracted DNA obtained from three yeast genera using Novozyme to lyse the cell walls differed significantly. When a set of universal primers were used in pure yeast cultures, PCR sensitivity for Saccharomyces exiguus was 2.5 pg (approx. 102 cells), for Candida mogii 12 pg (approx. 103 cells), and for Saccharomyces cerevisiae 20 pg (approx. 103 cells). In mixed yeast cell cultures, there was a 1000-fold (approx. 105~106 cells) decrease in sensitivity under the same PCR conditions.

Key words: polymerase chain reaction, universal primer, ribosomal RNA gene,yeast, internal transcribed spacer