COMPARISON OF DNA STANDARDS FOR REAL-TIME PCR-BASED QUANTIFICATION OF LACTOBACILLUS ACIDOPHILUS IN DAIRY PRODUCTS

Abstract

Probiotic bacteria are an essential part of the healthy gut microbiota. Fermented foods as potential sources of health-promoting bacteria can regulate the intestinal microbial population. However, the exact quantification of these bacteria in such multiple-strain matrixes continues to remain elusive. In this study, we evaluated the reliability of genomic DNAs and cloned recombinant plasmids as standard controls for absolute real-time PCR assay. The associated standard curves were constructed and used for the quantification of Lactobacillus acidophilus probiotics. All stages from the design and construction of standards and related curves met the criteria for high-quality products. There were no significant differences between the two enumeration methods. However, plasmid-based standard curves resulted in a lower detection limit than the curves of genomic DNA standards. Our findings showed that the non-linearized recombinant plasmids had long-term stability at high concentrations during storage at -20 °C, which strongly depended on the purification methods. We propose that the recombinant plasmid standards can supersede the traditional genomic DNA standards for accurate quantification of probiotic bacteria.