Development of cadmium specific recombinant biosensor and its application in milk samples

A novel and highly specific recombinant biosensor has been investigated for detection of highly toxic metal ion cadmium (Cd(II)) based on Escherichia coli DH5α (pNV12). The working principle of biosensor is based on the expression of gfp gene under the control of cad promoter and the cadC gene of Staphylococcus aureus plasmid pI258. Escherichia coli DH5α (pNV12) was tested for its response in the presence of different combinations of Pb(II), Cd(II) and Zn(II) ions. The investigation resulted in the development of a cadmium specific recombinant whole cell biosensor with a response time as low as 15 min. A good linear range for cadmium ion concentrations of 10-50 μg/l with R2 0.9946 was obtained and the detection limit of 10 μg/l. Further, we reduced the detection limit up to 5 μg/l by increasing the incubation time from 15 to 30 min. High throughput microarray techniques for testing of 48 samples simultaneously has also been developed with reaction volume miniaturized from 2000 μl to 200 μl. The developed biosensor was applied to detect cadmium ion concentration in milk samples collected from different areas of Punjab, India. Two samples collected from industrial areas, shown to have cadmium concentration above permissible limits (10 μg/l).