Q.Explain briefly the various processes of recombinant DNA technology.
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Imagine you have a cookbook with recipes from all over the world. Normally, you can only cook what's in one book at a time. But what if you could cut out the best recipe from one book and paste it into another, so your new book has a dish that no single cuisine ever had before? That is the basic idea behind Recombinant DNA Technology.
At its simplest, this technology is a way to take a piece of DNA (the genetic instruction manual) from one organism and join it with the DNA of a completely different organism. The result is a new, "recombinant" DNA molecule — a hybrid that never existed in nature. Think of it as genetic tailoring: you cut a gene from a human, a bacterium, or a plant, and stitch it into the DNA of another organism, often a bacterium or yeast. That host organism then reads the new instructions and starts producing the protein the inserted gene codes for.
Why does this matter? Because it lets us manufacture things that living organisms naturally make, but in a controlled, large-scale way. For example, the human insulin gene can be inserted into E. coli bacteria. These bacteria then become tiny factories, churning out human insulin that can be purified and given to diabetic patients. Before this technology, insulin had to be extracted from the pancreases of cows and pigs — a slow, expensive, and sometimes allergenic process.
The NCERT textbook (Class 12 Biology, Chapter 11) defines it precisely: Recombinant DNA Technology is the technique of joining DNA from two different species and inserting it into a host organism to produce a new genetic combination. The textbook highlights three key tools that make this possible:
- Restriction Enzymes – These are the "molecular scissors" that cut DNA at specific, predictable points. They allow scientists to cut out a desired gene cleanly.
- Vectors – These are the "delivery vehicles," usually plasmids (small circular DNA in bacteria) or viruses, that carry the foreign DNA into the host cell.
- Host Organisms – The living factory (like bacteria, yeast, or plant cells) that will replicate the recombinant DNA and produce the desired protein.
The core principle is genetic recombination — creating a DNA molecule that contains sequences from two or more different sources. This is not the same as natural reproduction or mutation; it is a deliberate, laboratory-made hybrid.
The process itself follows a clear sequence:
- Isolation of the desired gene (say, the human insulin gene) from the donor organism's DNA.
- Cutting both the gene and the vector DNA with the same restriction enzyme, creating matching "sticky ends."
- Ligation — using an enzyme called DNA ligase to permanently join the gene and the vector, forming the recombinant DNA.
- Transformation — inserting this recombinant DNA into a host cell (like a bacterium).
- Selection — identifying and growing only those host cells that successfully took up the recombinant DNA.
- Expression — getting the host cells to produce the desired protein in large quantities.
A common confusion is thinking this technology creates "new life." It does not. It creates a new genetic combination inside an existing living cell. The host organism remains the same species, but it now carries an extra instruction — like a factory that gets a new blueprint for a product it never made before. …
Recombinant DNA technology proceeds through a defined series of steps, starting with obtaining the DNA and ending with expression and recovery of the desired gene product. …
Recombinant DNA technology proceeds through isolation of DNA, cutting and joining DNA using enzymes, introducing it into a host, selecting successfully transformed cells, and expressing/harvesting the desired gene product.
Recombinant DNA (rDNA) technology involves altering the chemistry of genetic material (DNA/RNA) to introduce these into host organisms and change their phenotype. The main processes are:
1. Isolation of genetic material (DNA):
The cell/tissue is broken open (using lysozyme for bacteria, cellulase for plant cells, chitinase for fungi) to release DNA along with other macromolecules (RNA, proteins), which are then removed by treatment with RNase, protease, and purified by precipitation using chilled ethanol.
2. Cutting DNA at specific locations (restriction digestion):
Purified DNA is cut using restriction endonucleases, which recognise and cut DNA at specific palindromic nucleotide sequences, producing DNA fragments. The cut fragments are separated and visualised by gel electrophoresis and eluted from the gel for further use.
3. Amplification of the gene of interest (PCR):
The Polymerase Chain Reaction uses a thermostable DNA polymerase (Taq polymerase) and two primers to make multiple copies of the desired gene/DNA fragment in vitro, in a repeated cycle of denaturation, annealing and extension.
4. Ligation of DNA into a vector:
The amplified/cut gene of interest and a cut cloning vector (plasmid) are joined together using the enzyme DNA ligase, which forms phosphodiester bonds to create a recombinant DNA molecule.
5. Insertion of recombinant DNA into the host cell (transformation):
The recombinant DNA is introduced into a suitable host cell (e.g., bacterium) by making the host cell 'competent' to take up DNA — commonly by treating cells with divalent cations (e.g., CaCl2) followed by a heat shock, or by methods like micro-injection, biolistics ('gene gun'), or using disarmed pathogen vectors (e.g., Agrobacterium) for plant cells.
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Showing the 12 most recent of 18 on this concept.
- CBSE 2026Set 57/1/11 markMCQQ.Which of the following statements about plasmids is incorrect ? (A) Plasmids have the ability to replicate within the bacterial cell. (B) Their replication is controlled by chromosomal DNA. (C) They are autonomously replicating circular extra-chromosomal DNA. (D) They often carry antibiotic resistant genes.
›Reveal solutionSolution
The incorrect statement about plasmids is that their replication is controlled by chromosomal DNA; plasmids replicate autonomously.
Plasmids are fascinating and crucial components in the world of microbiology and biotechnology. They are essentially small, circular, extra-chromosomal DNA molecules found predominantly in bacteria, but also in some eukaryotes like yeast. Think of them as accessory genetic units that carry non-essential but often beneficial genes for the host cell.
Let's break down the given statements to understand which one is incorrect:
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Statement (A): Plasmids have the ability to replicate within the bacterial cell.
This statement is correct. A defining feature of plasmids is their ability to self-replicate. They possess their own origin of replication (ori) sequence, which allows them to initiate DNA synthesis independently of the main bacterial chromosome. This autonomous replication is what makes them incredibly useful as vectors in genetic engineering, as they can multiply along with the host cell, carrying the inserted foreign DNA.
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Statement (B): Their replication is controlled by chromosomal DNA.
This statement is incorrect. As mentioned above, plasmids replicate autonomously. This means their replication is independent of the bacterial cell's main chromosomal DNA replication. While the host cell's machinery (enzymes, nucleotides) is used for plasmid replication, the initiation and control of plasmid replication are governed by sequences within the plasmid itself, not by the bacterial chromosome. This independence is a key distinction between plasmids and the main chromosome.
ImportantThe autonomous replication of plasmids, independent of the host cell's chromosomal DNA, is a fundamental characteristic that distinguishes them and makes them invaluable tools in molecular biology.
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Statement (C): They are autonomously replicating circular extra-chromosomal DNA.
This statement is correct. This is a precise definition of a plasmid.
- Autonomously replicating: They have their own origin of replication and can replicate independently.
- Circular: Most plasmids are found in a closed circular form. …
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- CBSE 2026Set 57/2/11 markMCQQ.Assertion (A) : DNA Ligase is used to join DNA fragments. Reason (R) : It catalyses the formation of glycosidic bonds between nucleotides. (A) Both Assertion (A) and Reason (R) are true and Reason (R) is the correct explanation for Assertion (A). (B) Both Assertion (A) and Reason (R) are true, but Reason (R) is not correct explanation for Assertion (A). (C) Assertion (A) is true, Reason (R) is false. (D) Assertion (A) is false, Reason (R) is true.
›Reveal solutionSolution
DNA Ligase joins DNA fragments by catalysing phosphodiester bonds, not glycosidic bonds. So Assertion is true, Reason is false — option (C).
The question tests a very specific piece of molecular biology: what bond does DNA Ligase actually form? Many students memorise "DNA Ligase joins fragments" but forget the exact chemistry. Let's break it down cleanly.
Why the Assertion is correct
DNA Ligase is the enzyme used in DNA replication, repair, and genetic engineering to seal nicks in the DNA backbone. It joins two DNA fragments — for example, an Okazaki fragment to the growing lagging strand, or an insert into a plasmid vector. This is a standard fact in biotechnology and molecular biology.
Why the Reason is wrong
The Reason claims DNA Ligase catalyses the formation of glycosidic bonds between nucleotides. That is a critical error.
- A glycosidic bond links a sugar (deoxyribose) to a nitrogenous base (A, T, G, C) — that bond is formed during nucleotide synthesis, not by DNA Ligase.
- DNA Ligase catalyses the formation of a phosphodiester bond between the 3'-hydroxyl group of one nucleotide and the 5'-phosphate group of the next nucleotide. This is the bond that holds the sugar-phosphate backbone together.
Watch outA common mistake is confusing "joining nucleotides" with "forming the bond between sugar and base." The bond between nucleotides in a DNA strand is always phosphodiester, never glycosidic.
Step-by-step reasoning
- Evaluate Assertion (A): "DNA Ligase is used to join DNA fragments." …
- CBSE 2026Set ANNUAL1 markMCQQ.Given below are the steps carried out to construct a recombinant DNA.(i) Isolation of genetic material(ii) Insertion of recombinant DNA in the host cell /organism(iii) Obtaining the foreign gene product(iv) Amplification of gene of interest(v) Downstream processing. Which one of the following gives the correct sequences of these steps?(a)(i)(iii)(iv)(ii)(v)(b)(i)(iv)(ii)(iii)(v)(c)(ii)(i)(iii)(iv)(v)(d)(ii)(iv)(v)(iii) (i)
›Reveal solutionSolution
Constructing and using recombinant DNA follows a fixed logical sequence: first isolate and amplify the gene of interest, then insert the recombinant DNA into a host, let the host express the gene product, and finally process that product for use — i.e., (i) → (iv) → (ii) → (iii) → (v).
Recombinant DNA technology (genetic engineering) involves several sequential processes:
- (i) Isolation of genetic material (DNA) — the DNA must first be extracted in a pure form from the source cell, free of other macromolecules.
- (iv) Amplification of the gene of interest — using restriction enzymes to cut out the desired gene and, typically, PCR to amplify it, then ligating it into a suitable vector to form the recombinant DNA.
- (ii) Insertion of the recombinant DNA into the host cell/organism — the recombinant DNA (vector + gene of interest) is introduced into a competent host cell (transformation) and the host is then cultured/multiplied so the gene is expressed. …
- CBSE 2025Set ANNUAL1 markMCQQ.Which one of the following is not required for rDNA technology ?(i) DNA polymerase(ii) DNA ligase(iii) Restriction endonuclease(iv) Reverse transcriptase
›Reveal solutionSolution
Basic recombinant DNA technology always needs a cutting enzyme (restriction endonuclease), a joining enzyme (DNA ligase) and DNA-copying capability (DNA polymerase, e.g. for PCR); reverse transcriptase is only needed in the special case of starting from an RNA/mRNA template, so it is the one "not required" in general.
The core enzyme toolkit of rDNA technology includes:
- Restriction endonucleases: "molecular scissors" that cut DNA at specific recognition sequences to isolate the gene of interest and to open up the vector for insertion — always required.
- DNA ligase: joins ("glues") the cut gene of interest into the vector DNA to form the recombinant DNA molecule — always required.
- DNA polymerase: needed for amplifying DNA (e.g. via PCR) and is also the enzyme the host cell itself uses to replicate the recombinant plasmid — required in essentially every rDNA procedure. …
- CBSE 2024Set 57/3/11 markMCQQ.Which native plasmid did Stanley Cohen and Herbert Boyer use for the construction of the first recombinant DNA ? (A) Salmonella typhimurium (B) Streptococcus pneumoniae (C) Escherichia coli (D) Haemophilus influenzae
›Reveal solutionSolution
The first recombinant DNA experiment used a plasmid from Salmonella typhimurium — specifically, a small, naturally occurring plasmid called pSC101 — which was cut with a restriction enzyme and spliced with foreign DNA. The correct option is (A).
The key to this question is remembering the historical landmark experiment by Cohen and Boyer in 1973. They weren't just using any E. coli plasmid — they deliberately chose a plasmid from a different bacterial species to demonstrate that recombinant DNA could be constructed across species boundaries.
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Why not E. coli?
E. coli does have plasmids (like the famous pBR322, developed later), but Cohen and Boyer wanted to prove that DNA from one organism could be stably inserted into the plasmid of another. Using an E. coli plasmid would have been less convincing — it might have been argued that the foreign DNA was just recombining with native E. coli sequences. So they picked a plasmid from a different bacterium: Salmonella typhimurium.
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The actual plasmid: pSC101
This plasmid was isolated from Salmonella typhimurium by Stanley Cohen. It was small (about 9.4 kb), had a single EcoRI restriction site, and carried a tetracycline-resistance gene — perfect for selection. Boyer’s team used EcoRI to cut both pSC101 and a piece of foreign DNA (from a frog or another bacterium), then ligated them together. The resulting recombinant plasmid was introduced into E. coli, where it replicated and expressed the foreign DNA.
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Why the other options are wrong …
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- CBSE 2024Set ANNUAL1 markMCQQ.DNA or RNA segment tagged with radioactive molecule is called –(a) Probe(b) Vector(c) Clone(d) Plasmid
›Reveal solutionSolution
A radioactively labelled DNA/RNA segment used to identify a complementary target sequence is called a molecular hybridisation probe.
In recombinant DNA technology, a probe is a single-stranded DNA or RNA sequence tagged with a radioactive isotope (e.g., ³²P), used to detect the presence of a complementary nucleotide sequence in a sample through molecular hybridisation (e.g., in Southern blotting, or colony hybridisation for screening a clone carrying the gene of interest).
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- CBSE 2023Set TERM21 markMCQQ.A single strand of Nucleic acid tagged with a radioactive molecule is called:(a) Plasmid(b) Vector(c) Probe(d) Selectable marker
›Reveal solutionSolution
A single-stranded nucleic acid molecule tagged with a radioactive isotope, used to detect a complementary sequence, is called a probe.
In recombinant DNA technology, to confirm the presence of a specific gene or DNA sequence among many DNA fragments, a single-stranded DNA or RNA molecule complementary to that sequence is labelled with a radioactive isotope (commonly ³²P) — this labelled molecule is called a (radioactive) probe. It is used in techniques such as Southern/Northern hybridisation to specifically bind (hybridise) with its complementary sequence, which can then be detected by autoradiography. …
- CBSE 2023Set ANNUAL1 markMCQQ.The reagent that is used in the ELISA test is(a) polymerase(b) peroxidase(c) ligase(d) endonuclease
›Reveal solutionSolution
ELISA uses an enzyme such as peroxidase.
ELISA (Enzyme-Linked Immunosorbent Assay) detects an antigen or antibody by attaching an enzyme to a specific antibody. When the substrate is added, the enzyme catalyses a colour-producing reaction whose intensity indicates the amount of antigen present. Common reporter enzymes are horseradish peroxidase and alkaline phosph …
- CBSE 2022Set M1 markQ.What is the function of DNA ligase?
›Reveal solutionSolution
DNA ligase joins DNA fragments by sealing nicks with phosphodiester bonds.
DNA ligase catalyses the formation of a phosphodiester bond between adjacent nucleotides, thereby joining two DNA fragments (for example, joining a foreign DNA insert to a vector, or sealing Okazaki fragments during replication). I …
- CBSE 2022Set GO1 markQ.What is 'r' in rDNA?
›Reveal solutionSolution
'r' = recombinant; rDNA = recombinant DNA.
Concept. Recombinant DNA (rDNA) is an artificial DNA molecule created by combining a piece of foreign (donor) DNA with a vector DNA (such as a plasmid). The foreign gene is cut out with restriction endonucleases and joined to the cut vector by DNA ligase; the resulting hybrid molecule is the recombinant DNA. When i …
- CBSE 2019Set ANNUAL1 markQ.In what way Genomic DNA Library differ from C DNA Library?
›Reveal solutionSolution
A genomic library represents an organism's entire DNA including non-coding regions, while a cDNA library represents only the genes actually being expressed as mRNA in a specific cell type, without introns.
A genomic DNA library is a collection of clones that together represent the entire genomic DNA of an organism; it is prepared by isolating the whole genome, cutting it into fragments with restriction enzymes, and inserting these fragments into a vector — since it is made directly from genomic DNA, it includes both the coding sequences (exons) and the non-coding sequences (introns, regulatory regions, and so on). A cDNA (complementary DNA) library, in contrast, is prepared by first isolating mature mRNA from a particular cell or tissue and converting it into complementary DNA using the enzyme reverse transcriptase; because mature mRNA has already had its introns removed by RNA splicing, a cDNA library contains only exonic, protein-coding sequences and represents only the specific genes that were actively being expressed (transcribed) in that particular cell/tissue at the time …
- CBSE 2019Set ANNUAL1 markMCQQ.In certain cases for early and accurate detection of disease we may use-(a) ELISA(b) Culture(c) Chemical(d) Analytical
›Reveal solutionSolution
ELISA enables early and accurate detection of disease.
Early diagnosis of many diseases uses molecular/immunological techniques. ELISA (Enzyme-Linked Immunosorbent Assay) is based on antigen–antibody interaction and detects the antigen (pathogen) or antibodies against it, allowing detection even be …
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