Rapid and specific identification of medium-chain-length polyhydroxyalkanoate synthase gene by polymerase chain reaction

Abstract

A polymerase chain reaction (PCR) protocol was developed for the specific detection of genes coding for type II polyhydroxyalkanoate (PHA) synthases. The primer-pair, I-179L and I-179R, was based on the highly conserved sequences found in the coding regions of Pseudomonas phaC1 and phaC2 genes. Purified genomic DNA or lysate of colony suspension can serve equally well as the target sample for the PCR, thus affording a simple and rapid screening of phaC1/C2-containing microorganisms. Positive samples yield a specific 540-bp PCR product representing partial coding sequences of the phaC1/C2 genes. Using the PCR method, P. corrugata 388 was identified for the first time as a medium-chain-length (mcl)-PHA producer. Electron microscopic study and PHA isolation confirmed the production of mcl-PHA in P. corrugata 388. The mcl-PHA of this organism has a higher molecular weight than that of similar polymers produced by other pseudomonads.