Quantification of catechol dioxygenase gene expression in soil during degradation of 2,4-dichlorophenol

Abstract

The tfdC and C23O genes encode two catechol dioxygenases that catalyse ortho and meta cleavage of a key metabolite (chlorocatechol) of 2,4-dichlorophenol (2,4-DCP) metabolism, respectively. Primers were designed and a real-time PCR assay was developed to assess the abundance and expression of both tfdC and C23O genes in a soil amended with 2,4-DCP over a 21-day period. tfdC, the gene encoding the ortho cleaving dioxygenase, was significantly more abundant than the meta cleaving dioxygenase gene (C23O) throughout the experiment. The highest levels of tfdC were observed 2 days after amendment of soil with 2,4-DCP, at which stage the rate of 2,4-DCP degradation was at its maximum. In contrast, C230 copy numbers declined initially and peaked when degradation had slowed considerably. mRNA of the two chlorocatechol dioxygenase genes was not detected on day 0, but both genes were expressed after this time point. tfdC was expressed at a significantly higher level than C23O in 2,4-DCP-amended soil throughout the course of the microcosm, indicating the dominance of the ortho metabolic pathway. Phylogenetic analysis revealed a wide diversity of chlorocatechol dioxygenase genes in the 2,4-DCP-exposed soil examined.