Test: Tools of Recombinant DNA Technology - NEET MCQ

A restriction endonuclease is an enzyme that cuts the DNA molecule at, or near to, a specific nucleotide sequence to produce discrete DNA fragments that can be separated by gel electrophoresis.

Line in NCERT: "endonucleases make cuts at specific positions within the DNA."

Biolistics or gene gun is a direct or vectorless way used to introduce alien DNA into host cells. In this method of gene transfer, high velocity micro-particles of gold or tungsten, coated with DNA are bombarded on the plant cells. 

Line in NCERT: "cells are bombarded with high velocity micro-particles of gold or tungsten coated with DNA in a method known as biolistics or gene gun."

Escherichia coli and Agrobacterium tumefaciens are the microbes found to be very useful in genetic engineering. 
E. coli is a motile, gram negative, rod shaped bacterium which is a normal inhabitant of human colon. It is most extensively used in bacterial genetics and molecular biology.
Agrobacterium tumefaciens is a soil bacterium. It has Ti plasmid (Tumour inducing plasmid) and it can be used for the transfer of a desired gene in dicot plants.

Line in NCERT: "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. Similarly, retroviruses in animals have the ability to transform normal cells into cancerous cells." "When this DNA is transferred into Escherichia coli, a bacterium closely related to Salmonella, it could replicate using the new host's DNA polymerase enzyme and make multiple copies."

Gel electrophoresis is used to separate macromolecules like DNA, RNA and proteins. DNA fragments are separated according to their size. Proteins can be separated according to their size and their charge (different proteins have different charges).

Line in NCERT: "The DNA fragments separate (resolve) according to their size through sieving effect provided by the agarose gel. Hence, the smaller the fragment size, the farther it moves."

• The separated bands of DNA are cut out from the agarose gel and extracted from the gel piece.
• This step is known as elution.

Line in NCERT: "The separated bands of DNA are cut out from the agarose gel and extracted from the gel piece. This step is known as elution."

The origin of replication (ori) is crucial for initiating the replication of the DNA within the host cells, ensuring that the DNA is copied during cell division.

Ncert Topic: Cloning Vectors

Ncert line: Origin of replication (ori) : This is a sequence from where replication starts and any piece of DNA when linked to this sequence can be made to replicate within the host cells. This sequence is also responsible for controlling the copy number of the linked DNA

The drug resistance gene against penicillin is not typically used as a selectable marker in E. coli, as it does not provide a reliable means of selection compared to other antibiotic resistance genes.

Ncert Topic: Cloning Vectors ( Selectable Marker)

Ncert Line: Normally, the genes encoding resistance to antibiotics such as ampicillin, chloramphenicol, tetracycline or kanamycin, etc., are considered useful selectable markers for E. coli.

To evaluate the correctness of the statements:

- Statement i is correct; Hind II was indeed the first restriction endonuclease characterized and it cuts DNA at a specific six-base pair sequence.

- Statement ii is incorrect; while The convention for naming these enzymes is the first letter of the name comes from the genus and the second two letters come from the species of the prokaryotic cell from which they were isolated.

- Statement iii is incorrect; while exonucleases and endonucleases are both nucleases, not all endonucleases are restriction enzymes. Restriction enzymes specifically cut at defined sequences, while exonucleases remove nucleotides from the ends of DNA.

- Statement iv is correct; restriction endonucleases indeed recognize and cut specific palindromic sequences.

Thus, the correct options that reflect true statements are i, ii, and iv, which corresponds to Option C.

Ncert Topic:

Ncert Line:

The first restriction endonuclease–Hind II, whose functioning depended on a specific DNA nucleotide sequence was isolated and characterised five years later. It was found that Hind II always cut DNA molecules at a particular point by recognising a specific sequence of six base pairs.

The convention for naming these enzymes is the first letter of the name comes from the genus and the second two letters come from the species of the prokaryotic cell from which they were isolated.

Exonucleases remove nucleotides from the ends of the DNA whereas, endonucleases make cuts at specific positions within the DNA

Each restriction endonuclease recognises a specific palindromic nucleotide sequences in the DNA.

• Assertion (A) Analysis: The assertion is true because alternative selectable markers indeed facilitate a more efficient identification process for recombinant colonies, reducing the complexity associated with traditional antibiotic selection methods.
• Reason (R) Analysis: The reason is also true, as it accurately describes how these markers function by enabling visual differentiation through color change.
• Explanation: The reason directly explains the assertion, as the simplification of the identification process is a direct result of the color change method provided by alternative selectable markers.
• Ncert line: 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.