Biotechnology Test - 2

• Plasmids renature because they have supercoiled structures that have held the two strands of the helix together during denaturation.
• Chromosomal DNA is not able to renature, however, because its longer strands have become mixed with denatured proteins.

• Insertional inactivation is the inactivation of a gene upon insertion of another gene inside in its place or within its coding sequence.
• It help in identification of recombinant clones.

• Having two antibiotic resistance genes in the same plasmid helps in identifying transformants.
• Transformants take up the plasmid and also integrate it into recombinant DNA.

• In order to use the DNA for genetic engineering processes, it must be in pure form, free from other macromolecules.
• This is termed DNA isolation and is the pioneer step. Since the DNA is enclosed within the membranes, it is required to break open the cell to release DNA along with other macromolecules such as RNA, proteins, polysaccharides, lipids, etc. and from this mixture, DNA is purified.
• A release of DNA from a cell is achieved by treating the cells or tissues with enzymes such as lysozyme (bacteria), cellulose (plant cells), chitinase (fungus), etc.
• These enzymes degrade cell wall; plasma membrane degrading enzymes like lipase, etc., are also needed. Since yeast is a fungus and fungal cell wall is made of chitin (fungal cellulose), isolation of DNA necessarily requires the use of enzyme chitinase.

• The polymerase chain reaction is a technology in molecular biology used to amplify a single copy or a few copies of a piece of DNA.
• Calcium ions is not required in PCR technology.

• PCR is the short form for polymerase chain reaction which is the phenomenon which is used for amplification of the part-specific region of DNA as per our interest.
• The final step of PCR is extension, wherein Taq DNA polymerase (isolated from a thermophilic bacterium Thermus aquaticus) synthesises the DNA region between the primers, using dNTPs (denoxynucleoside triphosphates) and  Mg²⁺ .
• The primers are extended towards each other so that the optimum temperature for this
  polymerisation step is  72^∘ C. Taq polymerase remains active during high temperature induced denaturation of double stranded DNA.

In agarose gel electrophoresis, the DNA fragments separate out (resolve) according to their size or length because of the sieving property of agarose gel. It means, the smaller the fragment size, the farther it will move.

Restriction enzymes are also  called  'molecular scissors 'as they cleave DNA at or near specific recognition sequences  known as  restriction sites. These enzymes make  one  incision on each of the two strands of DNA and are also  called  restriction endonucleases.

• All the enzymes at higher temperature become inactive.
• Denaturation is done at high temperature as it leads to breaking down of hydrogen bonds between the bases of polynucleotides.

Genetic engineering involves:
(i) Identifying DNA with desirable genes.
(ii)  Introduction of identified DNA into the host.
(iii) Maintenance of introduced DNA in the host.
(iv)  Transfer of DNA in its progeny.

Hence, option C is correct.