Effect of Saliva Storage Conditions on Bacterial DNA Quantification by Real-time Polymerase Chain Reaction

Abstract

Objective: This research aimed to study the effects of temperature and duration of saliva samples storage on quantification of bacterial DNA by Real-time polymerase chain reaction (Realtime PCR) Materials and Methods: Human saliva samples were collected from 3 healthy volunteers, aliquoted into microcentrifuge tubes and stored in the following conditions; centrifuged immediately before storing at -80oC, kept at room temperature, on ice, or dry ice for 6 hours, 24 hours or 120 hours. Genomic DNA was extracted for quantitative analysis of total bacteria by Real-time PCR. Results: Absolute quantitative real-time PCR showed that concentrations of bacterial DNA from saliva samples stored at almost every condition which were stored at room temperature for 120 hours, on ice for every duration, or on dry ice for 6 or 24 hours were similar to that of the control, saliva centrifuged and frozen immediately after collection. However, bacterial DNA concentration from saliva stored at room temperature for 6 or 24 hours was significantly higher than that of the control. In contrast, the concentration of DNA from saliva stored on dry ice for 120 hours was significantly lower than that of the control. Conclusion: Temperature and duration had effects on bacterial DNA quantity. Saliva storage at room temperature within 24 hours had increasing bacterial DNA as quantify by Real-time PCR. Saliva storage on ice and dry ice could maintain bacterial DNA similar to sample centrifugation immediately and then frozen at -80oC. But long duration of storage, 120 hours, would decrease bacterial quantity.