SIMPLE DETECTION OF YERSINIA AND FRANCISELLA USING POLYMERASE CHAIN REACTION (PCR)

Background and Aim:Yersinia and Francisella are pathogens, known as a serious threat for public health. So, they are classifying as a biological agent category by Centers for Disease Control and Prevention (CDC). Laboratories (biosafety level 3) are required for working with these bacteria, but they are not easily available and also immunological and culture-based detection methods are complicated, so expanding molecular detection methods are valuable. Y. pestis causes plague and F. tularensis lead to tularemia. Due to lack of standard strain of these bacteria, designing a gene structure to use as positive control sample in detection tests is important.Methods:In this research, we used a synthetic construct, containing a conserved gene of each bacterium. Target region for Francisella is fopA and for Yesinia is caf1(F1 capsule antigen). After that, the construct inserted in PUC57 and transformed into E.coli DH5α. After an overnight culture, plasmids extracted and used for Monopelex PCR. Results was analyzed in 2% agarose gel.Results:We expect to see a 148bp band for Francisella and a 176 bp band for Yersinia. The results showed that amplification from each region was successful and expected bands were observed in electrophoresis.Conclusion:Due to lack of standard microbial strain for some bacteria, we can clone conserved regions of their genome into other bacteria and use them as positive control samples for other detection tests.

Keywords:

Francisella , Yesinia , PCR , Detection , Positive Control Sample