Read the given paragraph carefully and answer the following questions:
Since the DNA is enclosed within the membranes, we have to break the cell open to release DNA along with other macromolecules such as RNA, proteins, polysaccharides and also lipids. This can be achieved by treating the bacterial cells/plant or animal tissue with enzymes such as lysozyme (bacteria), cellulase (plant cells), chitinase (fungus). There are some basic steps in genetically modifying an organism – (i) identification of DNA with desirable genes; (ii) introduction of the identified DNA into the host; (iii) maintenance of introduced DNA in the host and transfer of the DNA to its progeny. Agrobacterium tumifaciens, a pathogen of several dicot plants is able to deliver a piece of DNA known as 'T-DNA' to transform normal plant cells into a tumor and direct these tumor cells to produce the chemicals required by the pathogen. Other molecules can be removed by appropriate treatments and purified DNA ultimately precipitates out after the addition of chilled ethanol. Selection of recombinants due to inactivation of antibiotics is a cumbersome procedure because it requires simultaneous plating on two plates having different antibiotics. 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. In this, a recombinant DNA is inserted within the coding sequence of an enzyme, β-galactosidase. This results into inactivation of the gene for synthesis of this enzyme, which is referred to as insertional inactivation.
Which one gives blue colored colonies if the plasmid in the bacteria does not have an insert?
Answer & explanation
Correct answer: option 3
The correct answer is Option (3) → The presence of chromogenic substrate
The plasmid carries the gene for the enzyme β-galactosidase. When the alien DNA is inserted inside this gene, the enzyme is not made (insertional inactivation). A chromogenic substrate is a colourless compound that the enzyme turns blue. So colonies whose plasmid has no insert make the enzyme and turn blue, while recombinant colonies stay colourless.
Option (1) The presence of cancerous cells — Incorrect. Cancer cells are animal cells that divide without control. They have nothing to do with selecting bacterial colonies on a plate.
Option (2) The presence of agarose — Incorrect. Agarose is the gel used in electrophoresis to separate DNA fragments.
Option (3) The presence of chromogenic substrate — Correct. Here the chromogenic substrate gives the colour. If the plasmid has no insert, the β-galactosidase gene is intact and the enzyme acts on the substrate, giving blue colonies; an insert inactivates the gene, so recombinant colonies stay colourless.
Option (4) The presence of ribonuclease — Incorrect. Ribonuclease is an enzyme that breaks down RNA. During DNA isolation it is used to remove the RNA mixed with the DNA. It plays no part in giving colour to colonies.
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 – “… Selection of recombinants due to inactivation of antibiotics is a cumbersome procedure because it requires simultaneous plating on two plates having different antibiotics. 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. In this, a recombinant DNA is inserted within the coding sequence of an enzyme, β-galactosidase. This results into inactivation of the gene for synthesis of this enzyme, which is referred to as insertional inactivation. The presence of a chromogenic substrate gives blue coloured colonies if the plasmid in the bacteria does not have an insert. Presence of insert results into insertional inactivation of the β-galactosidase gene and the colonies do not produce any colour, these are identified as recombinant colonies.”