Question

01. Lipases are one of the most important enzymes in detergent industry. in order to produce lipases in large scale, recombinant DNA technology can be used.

A .Outline the process of constructing a recombinant bacterium to produce lipase in large scale with all essential steps? and how you would identify a potential organism to isolate target gene foe lipase?(please mention the second bold question as "B" in your answer)

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Answer 1

A. Lipases are the enzymes that breakdown the fatty acids from the glycerol moiety in the lipid. The lipase encoding gene should be kept in front of a promoter in a plasmid which can be induced by IPTG or any inducible operon system. Then this plasmid needs to be transformed into the cells so that the gene can be expressed. A primary culture of this cells is made followed by the large secondary culture where a small inoculum of this primary culture is added and then induced by IPTG or respective inducer that activates the operon. By doing this the lipase enzyme will be expressed in a large scale and this helps in the large scale production of lipase by the recombinant microbe. The purification process can be carried out by affinity chromatography followed by dialysis.

B. The potential organism to isolate target gene of lipase will be by first knowing the gene sequence of the microbe that normally expresses the lipase enzyme. Then by using PCR, the lipase gene can be amplified and then inserted into the expression vector plasmid. In this way the potential organism can be found out for lipase gene extraction.

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