Persistence and renaturation efficiency of thermally treated waste recombinant DNA in defined aquatic microcosms

Abstract

To validate the possibility of horizontal gene transfer (HGT) from thermally denatured recombinant DNA discharged into the eco-system, a constructed plasmid was used to investigate the persistence and renaturation efficiency of thermally denatured recombinant DNA in defined aquatic microcosms. The results revealed that there was undecayed recombinant plasmid pMDLKJ material being discharged into the aquatic microcosms even after thermal treatment at either 100°C (using boiling water) or at 120°C (using an autoclave). The plasmid had a relatively long persistence time. At least 10² copies μL⁻¹ of a specific 245 bp fragment of the plasmid could be detected after 12 h and a specific 628 bp fragment could be detected up to 2 h. The thermally denatured recombinant DNA could efficiently renature and recover its functional double stranded structure in aquatic microcosms and the highest concentration of double-stranded DNA (dsDNA) occurred around 1 h after the thermally denatured DNA was added to the system. These results imply that when thermally treated recombinant DNAs are discharged into aquatic environments, they have enough time to renature and possibly transfer to other organisms. In addition, the recombinant DNA added to aquatic microcosms could be absorbed by the seston particles in water, such as mineral, organic and colloids particles with a maximum absorption value of about 5.18 ng L⁻¹. This absorbed DNA could persist longer in aquatic environments than free recombinant DNA, thus further favoring HGT.

Keywords:

• Waste recombinant DNA
• thermal treatment
• renaturation
• horizontal gene transfer (HGT)
• aquatic microcosm

Acknowledgments

We thank the anonymous referees for their constructive comments on the early version of this manuscript, which greatly improved the quality of our paper. This work was supported by China Natural Science Foundation (Grant no. 40571145 and 40871217) to Lei Wang, State Key Laboratory of Pollution Control and Resource Reuse Foundation (no. PCRRY11011), and the open fund of Shanghai Key Laboratory of Urbanization and Ecological Restoration (2010).