The utilization of RAPD-PCR for identifying thermophilic and mesophilic Bacillus species

A random amplified polymorphic DNA fingerprinting assay has been optimized that is able to discriminate between numerous thermophilic and mesophilic bacillus species and strains. Included in the analyses are thermophilic (able to grow at 55°C) strains of Bacillus stearothermophilus, B. kaustophilus, B. coagulans, B. sphaericus, B. thermodenitrificans, B. thermocatenulatus, B. thermoleovorans, B. licheniformis, B. brevis, B. thermoglucosidasius, B. caldolyticus, B. caldotenax, B. caldovelox, B. thermocloacae and B. smithii. Mesophilic strains of B. pumilus, B. subtilis, B. megaterium, B. circulans, B. cereus and B. mycoides can also be used for fingerprinting with the assay. Increasing the concentration of primer from 0.2 to 2.0 μM is shown to have a significant effect on increasing the number of amplification products that can be used for the discrimination or identification of individual strains or species. It is suggested that this may be a general way of improving the resolution of a RAPD protocol. The optimized conditions have been used successfully to trace B. stearothermophilus, B. licheniformis and other bacillus species and strains in an industrial setting.