Refer to the given diagram and answer the questions that follow : a) Which enzymes can be used for insertional

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Q. Refer to the given diagram and answer the questions that follow :

a) Which enzymes can be used for insertional inactivation of selectable marker genes?

(2)

b) Vector should not have more than one cloning site for the same restriction enzyme. Give reason.

(1)

c) What is the significance of rop?

(1)

d) Which region of the plasmid controls copy number? (1)

Karnataka PUCKarnataka II PUC Board 2026Subjective· 5mImportance★★★★★

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Figure — The stem refers to the pBR322 cloning-vector diagram and the answer reasons directly from its restriction site

The diagram is the cloning vector pBR322: insertional inactivation uses restriction sites within a resistance gene (e.g. BamHI/SalI in tetR, PstI/PvuI in ampR); a single cloning site per enzyme is needed to linearise the vector cleanly; rop controls plasmid replication; and ori controls the copy number.

The given diagram is the widely-used E. coli cloning vector pBR322, which carries two antibiotic-resistance (selectable marker) genes — ampicillin resistance (ampR) and tetracycline resistance (tetR) — an origin of replication (ori), the rop gene, and several unique restriction sites.

a) Enzymes for insertional inactivation of selectable marker genes (2 marks):

"Insertional inactivation" means inserting foreign DNA into a restriction site that lies within an antibiotic-resistance gene, which destroys (inactivates) that gene and lets us select recombinants.

Within the tetracycline-resistance (tetR) gene: recognition sites of BamHI and SalI (also HindIII). Inserting foreign DNA at BamHI or SalI inactivates tetracycline resistance, so recombinants become sensitive to tetracycline.
Within the ampicillin-resistance (ampR) gene: recognition sites of PstI and PvuI (and ScaI). Insertion here inactivates ampicillin resistance.

b) Why the vector should have only one cloning (recognition) site for a given restriction enzyme (1 mark):

So that the restriction enzyme cuts the vector at a single point, producing a single linearised molecule with defined ends into which the foreign DNA can be ligated. If there were more than one site for the same enzyme, the enzyme would cut the vector into several fragments, complicating gene cloning and preventing formation of a single stable recombinant DNA molecule.

c) Significance of rop (1 mark):

rop codes for the proteins involved in the replication of the plasmid — i.e. it regulates plasmid replication (and thus helps control copy number). …

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Same / Similar Concept — real previous-year questions on the same or a closely similar concept, not this exact question.

Showing the 12 most recent of 25 on this concept.

CBSE 2026Set A1 markMCQ
$Q. Which of the following is the sequence from where replication starts in cloning vector?$
(a) tet^R
(b) amp^R
(c) ori
(d) more than one
›Reveal solutionSolution

ori is the origin of replication; the correct option is (c).

A cloning vector must be able to replicate inside the host. The 'ori' (origin of replication) is the specific DNA sequence from which replication begins; any DNA linked to it can be made to replicate within the host cell. The ori also controls the copy number of the linked DNA.

…

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CBSE 2025Set ANNUAL1 markMCQ
$Q. Which one of the following is a genetic vector?$
(a) Phage
(b) Cosmid
(c) Plasmid
(d) Artificial chromosome
›Reveal solutionSolution

Plasmid is the most fundamental cloning/genetic vector; it is engineered with an origin of replication, selectable marker, and cloning site to carry foreign DNA into a host cell.

A cloning/genetic vector is a DNA molecule used as a vehicle to artificially carry a foreign DNA segment into a host cell, where it can be replicated and/or expressed. Plasmids are small, circular, double-stranded, extra-chromosomal DNA molecules found naturally in bacteria that replicate independently of the main bacterial chromosome. Engineered plasmids (e.g. pBR322) are the workhorse cloning vectors — they carry an origin of replication (ori), one or more selectable marker genes (e.g. antibiotic resistance), and unique restriction sites where foreign DNA is inserted.

…

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CBSE 2025Set ANNUAL1 mark
$Q. Why is it essential to have selectable marker in cloning vector?$
›Reveal solutionSolution

A selectable marker (e.g. an antibiotic-resistance gene) lets us identify and select only those host cells that have successfully taken up the vector, since transformation efficiency is naturally low.

When a recombinant DNA vector is introduced into host bacterial cells (transformation), only a small fraction of cells actually take it up — most cells remain untransformed. Since transformed and untransformed cells look identical, we need a way to selectively grow only the cells carrying the vector. This is achieved using a selectable marker gene on the vector — commonly a gene conferring resistance to an antibiotic (e.g. ampicillin resistance, tetracycline resistance) that the host bacterium is not normally resistant to. After transformation, cells are plated on medium containing that antibiotic: only cells that have taken up the vector (and hence the resistance gene) survive and grow, while untransformed cells are k …

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CBSE 2024Set E1 markMCQ
$Q. Which of the following is not a cloning vector for bacteria?$
(a) Bacteriophage
(b) Plasmid
(c) pBR322
(d) T-DNA
›Reveal solutionSolution

Plasmids, bacteriophages and pBR322 are cloning vectors used in bacteria; T-DNA is the transferred DNA of the Ti plasmid used to transform plant cells.

Cloning vectors for bacteria include plasmids (option b), bacteriophages (option a) and engineered plasmids such as pBR322 (option c). T-DNA (transfer DNA) is the portion of the Ti (tumour-inducing) plasmid of Agrobacterium tumefaciens that is trans …

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CBSE 2024Set E1 markMCQ
$Q. Which of the following is absent in pBR322?$
(a) Origin of replication
(b) Restriction site
(c) T-DNA
(d) Antibiotic resistant genes
›Reveal solutionSolution

pBR322 carries ori + restriction sites + antibiotic-resistance selectable markers; T-DNA is not part of it.

pBR322 is a well-known bacterial cloning vector. Its essential features are: an origin of replication (ori) that allows it to replicate inside the host (option a), several unique restriction sites where foreign DNA can be inserted (option b), and two antibiotic-resistance genes — am …

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CBSE 2024Set E1 markMCQ
$Q. If a foreign DNA is inserted in tetracycline resistant gene, the recombinant plasmid$
(a) Will lose ampicillin resistance
(b) Will lose tetracycline resistance
(c) Cloning would become easier
(d) Ampicillin resistance will become stronger
›Reveal solutionSolution

Inserting DNA into the tetR gene disrupts it — the recombinant plasmid loses tetracycline resistance (insertional inactivation).

In pBR322, foreign DNA is often inserted at a restriction site (like BamHI) lying inside the tetracycline-resistance (tetR) gene. This insertion disrupts the gene so it can no longer function — a process called insertional inactivation. As a result, cells carrying the recombinant plasmid become sensitive to tetracycline wh …

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CBSE 2024Set D1 mark
$Q. Match the following and write the correct pair. Column 'A':$
(i) Structural genes
(ii) T-Lymphocytes
(iii) Competition
(iv) PBR322
(v) Corona radiata. Column 'B':
(a) bone marrow
(b) vectors
(c) ovum
(d) detrimental interaction
(e) z, y and a
(f) RBCs. Find the correct match for item
(iv) 'PBR322' from Column 'B'.
›Reveal solutionSolution

'PBR322' pairs with option (b) 'vectors' — it is one of the earliest and most widely used cloning vectors.

In recombinant DNA technology, a vector is a DNA molecule (commonly a plasmid or bacteriophage) used as a vehicle to artificially carry a foreign piece of DNA into a host cell, where it can be replicated and/or expressed.

…

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CBSE 2024Set ZOOLOGY1 mark
$Q. Correct the underlined portion of the sentence: The cloning vector plasmids are found in monkeys (underlined).$
›Reveal solutionSolution

Plasmids used as cloning vectors are naturally occurring, self-replicating extra-chromosomal circular DNA found in bacteria (and some yeast), not in monkeys.

Plasmids are small, circular, double-stranded DNA molecules that exist independently of the bacterial chromosome and can replicate autonomously within a bacterial (or yeast) cell. Because they can carry a foreign DNA insert and replicate it along with themselves, plasmids (e. …

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CBSE 2023Set ANNUAL1 markMCQ
$Q. Plasmids are extra-chromosomal genetic material of which of the following?$
(a) Viruses
(b) Algae
(c) Fungi
(d) Bacteria
›Reveal solutionSolution

Plasmids are autonomously replicating circular DNA molecules found in bacteria, separate from the main bacterial chromosome, and they are the classic example of extra-chromosomal genetic material.

Bacterial cells (prokaryotes) contain one main circular chromosome, but many bacteria additionally carry one or more plasmids — small circular double-stranded DNA molecules that replicate independently of the chromosome. Plasmids often carry genes useful to the bacterium (e.g., antibiotic resistance) and are not essential for basic survival. Because they can be easily isolated, …

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CBSE 2023Set ANNUAL1 markMCQ
$Q. Which of the following is a cloning vector?$
(a) Plasmid
(b) RNA
(c) Mesosome
(d) Ribosome
›Reveal solutionSolution

A cloning vector is a DNA molecule that can carry a foreign gene into a host cell and replicate it there; plasmids are the classic and most widely used cloning vectors.

Recombinant DNA technology needs a 'vehicle' — a cloning vector — to carry a gene of interest into a host organism (commonly bacteria like E. coli) and enable its replication and, often, expression. Plasmids (e.g., pBR322) are ideal for this because they are small, circular, replicate independently of the host chromosome, and can be engineered with selectable markers (like …

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CBSE 2023Set ANNUAL1 mark
$Q. What is a plasmid?$
›Reveal solutionSolution

Plasmids are naturally occurring circular bacterial DNA molecules capable of independent replication, making them ideal vehicles (vectors) for carrying foreign genes.

A plasmid is a small, circular, double-stranded DNA molecule that exists separately from the main bacterial chromosome and can replicate independently of it, since it possesses its own origin of replication. Naturally occurring plasmids often carry genes conferring useful traits such as antibiotic resistance. In genetic engineering, plasmids (e.g., pBR322) are engineered and widely used as cloning vectors t …

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CBSE 2023Set ANNUAL1 markMCQ
$Q. In genetic engineering, antibiotics are used$
(a) in selecting healthy vectors
(b) as selectable markers
(c) to keep the culture free from infection
(d) both
(a) and (c)
›Reveal solutionSolution

In genetic engineering, antibiotics are used as selectable markers.

Cloning vectors carry antibiotic-resistance genes (e.g. amp^R, tet^R in pBR322). After transformation, cells are grown on a medium containing the antibiotic: only cells that have taken up the vector (and hence the resistance gene) survive. Thus the antibiotic-resistance gene works as a selectable m …

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Q.Refer to the given diagram and answer the questions that follow :

Concept understanding — Cloning Vectors and Selectable Markers (pBR322)

Cloning Vectors and Selectable Markers (pBR322)

Q.Refer to the given diagram and answer the questions that follow :

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