Development of an Absolute Quantitative Real-Time PCR (qPCR) for the Diagnosis of Aeromonas hydrophila Infections in Fish

Abstract

Keywords: Adhesin Gene; Molecular Diagnosis; Nile Tilapia; Septicemia Absolute quantitative real-time PCR (qPCR), a variation of poly-merase chain reaction (PCR), has been used to identify Aeromonas spp. in wastewater effluent and catfish [8,9]. Real-time qPCR allows DNA quantification, has high sensitivity and specificity and is accurate in detecting small numbers of bacteria. Use of qPCR also eliminates the need for gel electrophoresis. The fluorescent dye SYBR Green I used for this technique intercalates into double stranded DNA, remains stable at high temperatures, does not interfere with enzyme activity and binds to DNA molecules in a linear manner; SYBR Green I is also lower in cost compared with other intercalating agents on the market and is easy to use [10].