Abstract
Publisher Summary This chapter presents a procedure (i) to perform a polymerase chain reaction (PCR) amplification of the IS200 insertion sequence of Salmonella spp, (ii) to construct a graphite–epoxy composite (GEC) electrode, and (iii) to identify Salmonella spp . by using an electrochemical strategy based on GEC electrode. Amplification of the salmonella genome includes DNA extraction and PCR reaction. In DNA extraction, 1.5mL of an overnight culture of Salmonella enterica serovar Typhimurium ATCC 15038 on Luria–Bertani (LB) solid or liquid medium is centrifugated and the pellet is resuspended in 576 μL Milli-Q water. The PCR reaction is performed according to the kit manufacturer, in a 25 μL of reaction mixture containing PCR template, 200 μ mol/L of each deoxynucleotide triphosphate, 0.5 μ mol/L of each primer and 2.6 U of Taq polymerase, and in buffer containing 1.5 μ mol/L MgCl2. To identify Salmonella spp. by using an electrochemical strategy based on GEC electrode the amplified product coming directly from the PCR in 10×SSC is diluted. Further, the amplicon on GEC electrode is immobilized followed by denaturing, hybridization, and enzymatic labeling.
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