Molecular detection of Pseudomonas stutzeri by duplex-PCR technique

Abstract

Background and Objective: Duplex PCR is a widespread molecular biology technique that has the ability in specific and high sensitivity detection of microorganisms. This study was performed to evaluate the molecular identification of Pseudomonas stutzeri using duplex PCR. Methods: In this descriptive-laboratory study, Pseudomonas stutzeri ATCC 17588 bacteria was purchased from genetic resources center and after culturing the bacteria, DNA was extracted in the exponential growth phase using boiling method. Duplex PCR was carried out for specific identification of the bacteria subsequently. The primers were designed using catA and nirP gene sequences. Sensitivity and specificity of duplex PCR technique were investigated using 5 bacteria. Results: The amplification of two bands of 512 bp and 249 bp for catA and nirP genes were observed, respectively. The specificity was 100% .The sensitivity of 0.048 ng/µL of genomic DNA was determined for catA and nirP genes, respectively. Conclusion: Duplex-PCR molecular method with its sensitivity and proper feature and high potential for identification of Pseudomonas bacteria can be applied as a routine method in well-equipped laboratories by expert technician to identify suspicious cases.

Keywords

• Pseudomonas stutzeri
• Duplex-PCR
• catA gene
• nirP gene