Construction And Preliminary Application Of Bio-engineered Bacteria For Detection Of Environmental Cadmium

ObjectiveA new method was established to detect environmental cadmium ,in which green fluorescent protein gene(gfp) and cadmium-specific promoter PcadA were ligated to form a recombinant plasmid.MethodsCadmium-specific promoter PcadA was obtained by PCR from pPcadA and was ligated to E.coli-B.subtilis shuttle vector pNGFP with gfp being reporter gene ,after which the recombinant plasmid was transformed into 1A751 strain to form the bio-engineered bacteria.To assess the selectivity of the bio-engineered bacteria as a metal-ion biosensor, the 1A751 strain containing plasmid pPcadA-gfp was tested in the presence of various heavy metal ions respectively, and the fluorescence intensity was measured .To estimate the biosensing ability, the strain was induced by Cd2+ of varied concentrations, and the response was observed after different exposure time.ResultsAfter PCR and DNA sequence analysis, the targeted DNA was successfully inserted into the vector and was expressed as anticipated.The strain responded obviously to the cadmium cations, and no induction was observed as to other metal ions(As3+,Co2+,Cu2+,Fe2+,Hg2+,Mn2+,Ni2+,Pb2+,Sb3+,Sn2+,Zn2+).The data confirmed that the strain as a biosensor had a sound specificity for detection of cadmium cations.After different exposure time, the strain was induced by varied concentrations of Cd2+.The data indicated that it was sensitive to Cd2+ within the detection range of 1μM -500μM,and the fluorescence intensity was positively correlated to the cadmium ionic concentrations within 1μM to 250μM. The detection limit was 1μM,and the response time was about 15 minutes. After exposing to Cd2+ 15 minutes to 10 hours, the strain still had a good response to it within the detection range.ConclusionBy using the bio-engineered construct, a novel cadmium detection method is expected to be established.