Clone and Analysis of Atrazine Chlorohydrolase Gene of Arthrobacter sp. T3AB1 Isolated from Northeast China

Atrazine is a triazine herbicide widely used throughout the world in crop fields but it can cause long residual effects such as a phytotoxic response to the subsequent crop. Therefore, it is an urgent issue to effectively control and solve the contaminant of atrazine regularly in rain, ground water and crop fields. In this study, we reported one bacterial strain of Arthrobacter sp. T3AB1. It was capable of utilizing Atrazine as the sole nitrogen and carbon source isolated from the corn field in Nehe (Heilongjiang, China) where had seriously suffered the phytotoxicity of atrazine. The exoenzyme and endoenzyme extracted from T3AB1 and cell debris were all assayed for enzyme activities and the critical genes that could degrade atrazine was cloned by PCR; the specific expression of trzN gene was analyzed by Quantitative Real-time PCR (qRT-PCR). Our results indicated that the endoenzyme showed higher activities in a pH 6.0 phosphate buffer and could degrade 93.61% of atrazine. sequence analysis of the trzN, atzB and atzC genes demonstrated that these genes had very highly conserved domains. We also obtain the full length of trzN gene (1371-bp in length, accession no. GU459314). Compared to the reported hydrolases, seven mutation sites were found in this gene: 67F/F/Y→Y, 117E/Q/E→E, 197E/E/E→H, 305W/W/W→S, 299P/S/P/→P, 342A/A/A→T and 331D/D/D→E. QRT-PCR suggested that the trzN gene expression of T3AB1 strain cultured in 100 mg/L MSM+AT(Mineral salt medium including Atrazine) for 36 h was up-regulated, nearly twice as much as that in LB medium at the same time. It will be the molecular evidence and lay the foundation for the research of atrazine catabolic pathway by T3AB1 in the future.