Question

2) With reference to production of recombinant DNA answer the following questions: i) what is "plasmid" and how is it isolated from a cell (1 point); ii) what is "insert" (1/2 point); iii) name the enzyme used for cleaving and generating sticky ends on the plasmid and insert (1 point); iv) name the enzyme used for joining sticky ends of plasmid and insert to produce a molecule of recombinant DNA (1 point); v) name the most common microbial host cell used in the replication of recombinant DNA (172 point); vi) name two important proteins synthesized using recombinant DNA technology (1/2 point each)?

Answer 1

Answer 1. Plasmid is found in bacteria and it is an extra-chromosomal DNA molecule that can replicate independently.

Plasmid isolation is important step of recombinant DNA technique which is obtain from the method of alkaline lysis by breaking the cell of bacteria. Alkaline lysis buffer made up of detergent sodium dodecyl sulfate and sodium hydroxide. The detergent is used to break the phospholipid bilayer membrane and to denature the protein of cell membrane alkaline is used. Plasmid isolation involved many step like agitation, centrifugation, precipitation and removal of cellular debris before isolation of plasmid.

Answer 2. Insert is the introduction of recombinant DNA molecule in the host organism to produce desired protein.

Answer 3. Enzyme restriction endonuclease cleaves DNA at specific site and produce fragment of DNA.

Answer 4. DNA ligase is the enzyme that is used to join the DNA strand by making phosphodiester bonds. It ligate the desired DNA fragment at target site of vector.

Answer 5.Most common microbial host cell for the purpose of DNA recombinant technique is bacteria Escherichia. Coli.

Answer 6. Important protein synthesized by recombinant DNA technology is insulin and human growth hormone.