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Volume 6, Issue 4 (Nov 2018)                   Res Mol Med (RMM) 2018, 6(4): 0-0 | Back to browse issues page

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Pourmahdi N, Zeinoddini M, Dehghan Esmatabadi M J, Sheikhi F. Simple and Rapid Detection of Yersinia pestis and Francisella Tularensis using Multiplex-PCR. Res Mol Med (RMM). 2018; 6 (4)
URL: http://rmm.mazums.ac.ir/article-1-293-en.html
Abstract:   (119 Views)
Background: Yersinia pestis causes plague and Francisella tularensis lead to tularemia, which are known as newborn and retire diseases. Immunological and culture-based detection methods of these bacteria are time-consuming, costly and complicated. On the other hand working on these bacteria are required advanced equipment laboratories and the genome isolation methods are difficult. Therefore, design and synthesis of a gene structure as positive control is a smart strategy for molecular detection of these bacteria.
Materials and method: We used a synthetic construct, containing a conserved gene of Francisella (fopA) and Yersinia (caf1). Conserved regions of each gene were determined and a construct with both genes was designed and used for multipelex PCR assay. The sensitivity of this assay examined by serial dilution of extracted plasmid and the specificity of method examined using genome of   E.coli, Salmonella, Enterobacter, Vibrio as template. PCR products were analyzed in agarose gel electrophoresis.
Results: The data was showed a clear dual band in the regions of 107bp and 176bp that confirming the fopA and caf1 genes, also another 351bp band has been seen which is the reason of amplification by forward primer of fopA and reverse primer of caf1 which after optimization of assay this target amplification had reduced. The sensitivity of this assay was determined about 36×10 -3 ng/µl and the selectivity test confirmed the suitable specificity of this method for the detection of target genes.
Conclusion: This multiplex PCR technique could be used in research laboratories in order to identification of these important pathogens.
     
Type of Study: Research | Subject: Molecular biology
Received: 2019/03/9 | Accepted: 2019/05/8

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