ABSTRACT
Fast and accurate detection of Salmonella typhimurium (S. typhimurium) is needed. This study aims to design and test the primer optimum temperature for the S. typhimurium flagellin gene (fljB) with the Gradient Polymerase Chain Reaction (PCR) technique. The in silico analysis using Professional Clone Manager program 9.2 for the primer design showed that the fljB primer pair could amplify S. typhimurium fljB fragments to produce amplicon with size 106 base pairs (bp). Evaluation of the primer ability of the design result and determination of the optimum temperature to produce amplicons of the right size was carried out by Polymerase Chain Reaction (PCR) Gradient. The temperature variation on PCR-Gradient is 56–61°C. The S. typhimurium isolate as a template for the Gradient PCR process has a concentration of 53.75 ng/µΛ with purity (A260/280) of 1.85. The results of the study generously information that the fljB primer provides optimal results at annealing temperatures 60°C. This is indicated by the production of a DNA band with a size of 106 base pairs in accordance with the analysis in silico and has a thickness band intensity. Based on the results obtained, it can be concluded that the optimal temperature of the fljB primer of S. typhimurium is 60°C. Furthermore, the primers and optimum temperatures obtained were used for the Real Time PCR stage in the development of sensitive, specific and faster methods of detection of food poisoning bacteria.
