Cloning Vectors
Read the following passage carefully and answer the given questions.
In order to link the alien DNA, the vector needs to have very few, preferably single, recognition sites for the commonly used restriction enzymes. The ligation of alien DNA is carried out at a restriction site present in one of the two antibiotic resistance genes. For example, you can ligate a foreign DNA at the BamH I site of tetracycline resistance gene in the vector pBR322. The recombinant plasmids will lose tetracycline resistance due to insertion of foreign DNA but can still be selected out from non-recombinant ones by plating the transformants on ampicillin containing medium. The transformants growing on ampicillin containing medium are then transferred on a medium containing tetracycline. The recombinants will grow in ampicillin containing medium but not on that containing tetracycline. But, non-recombinants will grow on the medium containing both the antibiotics. In this case, one antibiotic resistance gene helps in selecting the transformants, whereas the other antibiotic resistance gene gets ‘inactivated due to insertion’ of alien DNA, and helps in selection of recombinants. Therefore, alternative selectable markers have been developed which differentiate recombinants from non-recombinants on the basis of their ability to produce colour in the presence of a chromogenic substrate.
The selectable marker, which differentiate recombinants from non-recombinants on the basis of ability to produce colour, is due to -
Answer & explanation
Correct answer: option 3
The correct answer is Option (3) → Insertional inactivation
(1) Antibiotic resistance genes — Incorrect. These are the older selectable markers. They need the cells to be plated on two plates with different antibiotics, and they do not give any colour.
(2) Insertional activation — Incorrect. This is not a term used for markers. Insertion of the alien DNA switches the gene off, not on.
(3) Insertional inactivation — Correct. The alien DNA is inserted inside the coding sequence of an enzyme, β-galactosidase. This inactivates the gene for this enzyme. With a chromogenic substrate, colonies without an insert are blue, and colonies with an insert stay colourless. So the recombinants are told apart by colour.
(4) Non-insertional inactivation — Incorrect. This is not a real term.
Note: The passage is taken from NCERT. NTA has put lines of the NCERT text together to make it. Most CUET (UG) questions come straight from NCERT, so study the book line by line, including figures and their titles.
NCERT Extract:
Cloning Vectors – …
