A Study of the Stability of Escherichia coli on DNA Banking Card (DBC) for Molecular Tests

Objective: Traditional methods for storing microbial DNA samples use the low-temperatures of ۴ °C, -۲۰°C or -۸۰ °C. The DBC Card is a new method of storing microbial DNA at room temperature. This study evaluates the stability and determine the best storing method of Bacterial DNA on the DBC card. Methods: In this study, we used a pair of primers from the conserved domain of ۱۶srRNA designed to identify Escherichia coli. Four different types of samples we prepared: i) bacterial suspension, ii) bacterial DNA extracted by the phenol – chloroform method, iii) bacterial lysate with lysis buffer and iv) bacterial DNA produced by the boiling method. All four samples were spotted on separate DBC cards and dried at room temperature. After periods of ۳, ۵ and ۷ months, Escherichia coli samples were checked for DNA stability with two molecular techniques, conventional PCR with ۱, ۲ and ۳ disks as a source of bacterial DNA (۱ mm diameter) and Real-Time PCR. Results: Data showed that DNA stability was maintained after ۷ months on a DBC disk, even using only one disk as a DNA source. The bacterial suspension was the best method for long-term storage of Escherichia coli DNA on the DBC Card. Conclusion: In traditional methods for storing sample, DNA quality reduces after freezing and thawing. However in the DBC method, DNA quality was maintained for a long duration. Advantages of the DBC method are easy sample handing, low cost, faster extraction, and reduced individual and environmental contamination.

کلیدواژه ها:

Real-Time PCR ، Real-Time PCR ، DNA Banking Card (DBC) ، Bacteria Suspension ، Storing Method of Bacterial DNA ، سوسپانسیون باکتری ، کارت نگهداری DNA ، روشهای نگهداری DNA باکتری