Cloning and Characterization of Cobalt and Zinc Tolerance Genes by Using Metatranscriptomics Approach

Abstract

Heavy metals are probably toxic to the living beings depending upon their concentration and sensitivity in the ecosystem. These living beings starting from unicellular to multicellular and prokaryotes to eukaryotes have developed various known and unknown mechanisms to adapt such stressed or adverse environment. Functional environmental transcriptomics plays a significant role in recognizing genes responsible or part of these unknown yet important pathways. Metatranscriptomics approach provides information about expressed subset of genes within a microbial community. In the present study, functional metatranscriptomics approach was adopted to identify cobalt tolerance genes expressed by eukaryotic species. Size fractionated libraries constructed in the modified pFL61 yeast- E.coli shuttle vector screened through yeast transformation for the presence of cobalt and zinc tolerant cDNAs followed by functional complementation assay for the resistant clones. In this study cobalt sensitive Δcot1 mutant strain and zinc tolerant Δzrc1derived from wild strain BY4741 was used. Yeast transformants showing cobalt and zinc tolerance were identified by plating on medium supplemented with respective metal and characterized by studying their growth kinetics.