Design of Oligonucleotide Primers for Amplification of the tolC Genes of Achromobacter insolitus LCu2

The RND-type efflux systems are the key components providing for resistance of gram-negative bacteria to antibiotics and heavy metals. The present work reports the design of specific oligonucleotide primers for the genes encoding the TolC outer membrane proteins, the RND systems components, and the housekeeping genes (gyrB, nusA, lepA, rpoB) for Achromobacter insolitus strain LCu2. Various bioinformatic tools were used to design the oligonucleotide primer pairs with the optimal characteristics: amplicon length 100–300 bp; melting temperature ~60°C; and G+C composition 50–60%. Experimental validation by electrophoresis and melting curve analysis data confirmed the specificity of oligonucleotide primer pairs, which indicated their applicability for further use in investigation of the tolC genes expression by real-time polymerase chain reaction.