Journal of Environmental Biology

Application of quantitative real-time PCR for quantification of Rhodococcus sp. EH831 in a polyurethane biofilter

 

E.H. Lee¹, K.S. Cho*¹ and H.W. Ryu*²

¹Department of Environmental Science and Engineering, Ewha Womans University, Seoul, 120-750, Republic of Korea

²Department of Chemical and Environmental Engineering, Soongsil University, Seoul, 156-743, Republic of Korea

(Received: March 07, 2008; Revised received: June 25, 2008; Accepted: July 15, 2008)

Abstract: Rhodococcus sp. EH831 is a microbial species that can degrade volatile organic compounds. We optimized a method for monitoring quantitative real-time PCR (qRT-PCR) of EH831 that was incorporated into a polyurethane (PU) biofilter. When the genomic DNA of EH831 was directly extracted from a PU sample with immobilized EH831, the recovery efficiency was very low due to DNA absorption into the PU. DNA amplification during PCR was also inhibited by PU impurities. Therefore, a pre-treatment step was necessary. We successfully recovered cells from the PU by squeezing the matrix, adding sterilized water, and vortexing. The recovery efficiency ranged from 105 to 144%, and there was no statistically significant difference. We designed a novel TaqMan probe for EH831 and demonstrated its high specificity for EH831. The detection range for EH831 was 10⁵-10¹¹ cfu ml^-1. The method described in this study can be used to investigate the relationship between quantitative analysis of Rhodococcus sp. EH831 and PU biofilter performance.

Key words:? Biofilter, Rhodococcus sp., Polyurethane, Quantitative real-time PCR, Monitoring

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