Development of lead (Pb) tolerant strain by protoplast technology and their remediation.

The lead is poisonous metal and because of its chemical nature it acts as an environmental contaminant through the water or soil and it becomes toxic to humans. The toxicity of Pb occurs as a change in the conformation of nucleic acid and protein, inhibition of enzyme activity, disruption of membrane function and oxidative phosphorylation. For protoplast preparation, the removal of the cell wall and protoplast formation obtained by specific lytic enzyme. In cytoplasmic membrane, the envelope of bacteria consists of overlying cell wall. From hypertonic environment, the complete cell wall removal occurs due to which it maintains the osmotic integrity of the cell and produces the protoplast. In current work, protoplasts were produced by specific lytic enzyme (lysozyme and macerozyme), chemo fused (with the help of Polyethylene Glycol) and regenerated from strains Staphylococcus sp. and Bacillus sp. The fused protoplast was spherical in shape observed under microscopy. Colonies were screened on specific medium supplemented with Pb (Concentration at the rate of 1.5mM). One resistant colony (MICBT-1) was selected and further examined and applied for the transformation of Pb in the broth medium. The strain removed 98% of Pb at 1mM concentration. Next, sucrose containing medium was best which gives maximum protoplast regeneration. From various organisms, fusion technique has been used to combine the genes to create the strains having desired properties. This is a significant technique for engineering of bacterial strains for advantageous applied properties. Further MICBT-1 applied in artificially contaminated soil and removed maximally in exchangeable fraction (remains up to 0.05 mM). An efficient bioremediating agent for lead transformation from soil and water is expected to ease the ever-increasing problem. Further, it is needful to obtain new strain with the help of protoplast technology which can reduce the pollutant. This lead tolerant strain can be applied for bioremediation purposes in the Pb contaminated soil and water environment.