Bioremediation of mineral mercury via construction of recombinant vector pET21a(+)-merA

Subject Areas : Microbial Biotechnology

Hamideh  Baghi Sefidan ¹ (M.Sc. student, Department of Agricultural Biotechnology, College of Agriculture, Azarbaijan Shahid Madani University, Tabriz, Iran.)
Alireza  Tarinejad ² (Associate Professor, Department of Agricultural Biotechnology, College of Agriculture, Azarbaijan Shahid Madani University, Tabriz, Iran.)

Received: 2016-10-22 Accepted : 2017-01-10 Published : 2017-05-22

Keywords: Mercury, Bioremediation, merA gene, recombinant vector,

Abstract :

Background & Objectives: Inorganic mercury entrance into the environment through industry and agriculture is one of the most serious environmental hazards in the country. Microbial bioremediation is considered as one of the practical solutions to clean up pollutant ions from the environment. The present study was conducted to construct a recombinant vector including merA gene, and also to investigate its expression in order to clean up mercury. Material & Methods: merA gene from mercury resistance bacterial genome was isolated, and subsequently cloned into the pET21a(+) expression vector. Confirmation of cloning target gene was achieved by PCR and restriction enzyme digestion. Then pET21a(+)-merA recombinant vector was cloned into E.coli strain BL21. In order to assess increased resistance to mercury in recombinant bacteria, and the functionality of the enzyme produced by the recombinant vector, the growth rate of recombinant BL21 E. coli strains to contain merA gene, and without it was measured in a medium containing mercury for 48 hours. Results: The results showed that the growth of E. coli strains without merA gene was strongly affected after introducing mercury into the media till 12 hours, and bacteria would not be able to grow at 10 ppm and 20 ppm mercury levels. However, transformed bacteria with pET21a(+)-merA vector showed suitable growth in mercury-containing media. SDS-PAGE analysis of recombinant bacterial proteins on acrylamide gel showed the highest MerA (62KDa) expression following 16 hours induction with 1mM IPTG at 37ºC. Conclusion: Overall, the growth ability of merA- containing recombinant bacteria reflects the action of MerA protein in transformed bacteria. Furthermore, increasing the resistance of recombinant bacteria to the mercury indicates that environmental heavy metal pollutants can be cleaned up properly through the construction of recombinant vectors.