关键词: 扩展青霉, 基因组DNA, PCR, 优化

Abstract: The genome DNAs of Penicillium expansum and 6 control fungi were extracted by glass bead method, benzyl chloride method and their combination, respectively. The concentration and purity of DNAs were analyzed by UV spectroscopy and gel electrophoresis. Meanwhile, a pair of primers with a size of 288 bp were designed and synthesized based on a conservative sequence of Penicillium expansum's polygalacturonase gene to conduct the PCR detection. The results showed that combined use of glass bead and benzyl chloride was more effective than either alone. Good specific amplification of DNA was obtained, and the optimal range of annealing temperature was between 53 ℃ and 59 ℃, and the optimal range of primer concentration between 0.04 μmol/L and 0.16μmol/L, and the optimal range of template concentration between 2.40μg/mL and 5.28μg/mL. Meanwhile, dNTPs concentration had little influence on PCR amplification. The method requiring only 3 to 4 hours could evidently enhance detection efficiency when compared to traditional methods. As a result, it has promising potential to be further applied in practice.

Key words: Penicillium expansum, genome DNA, polymerase chain reaction (PCR), optimization