A Novel qPCR Method for the Detection of Lactic Acid Bacteria in Fermented Milk

Abstract

The number of live lactic acid bacteria (LAB) is an important quality indicator for yogurt, the quantitative testing of LAB has become an important task in the evaluation of product quality and function. By analyzing and comparing the performance of 16S rRNA gene and tuf gene used in absolute quantification, the tuf gene with copy number 1 was selected as the target gene of six LAB. By drawing a standard curve to achieve qualitative and quantitative detection of six strains of LAB, the detection range was found to be 1 × 103–1 × 108 copies/µL. The traditional plate colony count and Flow Cytometry (FCM) were compared with the method of qPCR, which was used in this experiment. Meanwhile, the confocal laser microscope combined with STYO 9 and propidium iodide dyes was used to determine that the content of viable bacteria in the yogurt was more than 90%, which proved that the detection result using qPCR method was closer to the true level of LAB in yogurt. Compared with the existing methods, the method in this study allowed the qualitative and quantitative detection of the six kinds of LAB in yogurt, and the distribution of live and dead bacteria in yogurt could be calculated.

Keywords

• qPCR
• lactic acid bacteria
• yogurt
• laser confocal microscope
• <i>tuf</i> gene
• flow cytometry