Genus-specific Primer Design for Molecular Identification of Rhodobacter sp. Origin of the 16S rRNA Gene

Abstract

Rhodobacter is one of the purple phototropic bacteria widely used in aquaculture activity. It is particularly difficult in identifying and distinguishing Rhodobacter from other non-sulfur purple bacteria. This study aimed to evaluate primer pairs designed based on the 16S rRNA gene from Rhodobacter as an effective and efficient molecular identification that can be performed in the laboratory using conventional PCR techniques. The 16S rRNA sequences from Rhodobacter and other bacteria were aligned and analyzed for the high-variation region to capture the desired target amplicon. Forward and reverse primer pairs were designed based on the appropriate target amplicon. The primer pairs were also assessed for their feasibility based on ten assessment criteria for the quality of the PCR primers. The PCR optimization results obtained showed a specific band with a size of 548 bp according to the amplicon target size in the Rhodobacter samples tested and did not show any cross-reaction with other comparator bacteria, and no dimers or other non-specific bands were found. Based on the results obtained, it can be concluded that the primer pairs designed in terms of quality and specificity are good and can be used as alternative specific primers for the genus Rhodobacter sp.