Ncert Solutions Biotechnology Principles And Processes Class 12 Notes & Important Questions

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Ncert Solutions Biotechnology Principles And Processes Class 12 Notes


Class12th 
Chapter No11
ProvidingQuestions And Answers, Notes & Numericals PDF
Chapter NameBiotechnology: Principles and Processes
BoardCBSE
Book NCERT
SubjectBiology
Medium English
Study MaterialsFree VVI Study Materials are Available
Download PDF Biotechnology principles and processes ncert pdf

Biotechnology principles and processes class 12 notes


1. Biotechnology in the modern concept, refers to the processes which use genetically modified organisms to make various commercial products on a large scale.

Ncert Solutions Biotechnology Principles And Processes Class 12 Notes Important Questions

As opposed to the earlier concept that could mean making curd, bread or wine, which are all microbe-mediated processes. The European Federation of Biotechnology (EFB) has given a definition of biotechnology as, the integration of natural science and organisms, cells, parts thereof and molecular analogues for the products and services’. Examples of modern biotechnology are

(i) in vitro fertilisation to make a ‘test-tube’ baby.

(ii) synthesising a gene and using it.

(iii) developing a DNA vaccine.

(iv) correcting a defective gene (gene therapy). 

2. Principles of Biotechnology

The two core techniques of modern biotechnology are 

(i) genetic engineering to create a modified DNA. 

(ii) growing genetically engineered organisms to produce the desired product.

3. Genetic engineering is a deliberate modification of an organism’s DNA, using various techniques. This altered DNA (recombinant DNA) is then introduced into the same or different organism to change its phenotype. This is followed by growing this genetically modified microbe/eukaryotic cell in large quantities, maintaining a sterile (microbial contamination-free) environment, for the manufacture of biotechnological products like antibiotics, vaccines, enzymes, etc. 

(1) Traditional hybridisation procedures used in plant and animal breeding, often result in the inclusion and multiplication of undesirable genes along with the desired ones.

(ii) Genetic engineering overcomes this problem as it creates a recombinant DNA, by introducing only one or few desirable genes into the target organism.

The technique of genetic engineering includes 

(a) the construction of recombinant DNA.

(b) gene cloning.

(c) gene transfer. to no butanol

4. Making a genetically engineered product using recombinant DNA technology involves the following steps:

(1) Collecting a DNA (or chromosome) sample from a desired organism. 

(ii) Cutting this DNA into smaller fragments. 

(iii) Isolating (separating) the gene of interest from among these fragments. 

(iv) Combining it with the DNA of a vector (an agent that will carry it into the desired host).

(v) Introduction/injection/transfer of this DNA in a desired host organism. 

(vi) Integration of this DNA in the host genome. 

(vii) Successful replication or cloning (to make more copies of recombined DNA).

(viii) Maintenance of introduced DNA in the host and ensuring transfer of the DNA to its progeny. 

(ix) Its successful expression in a host organism so that the required change is expressed in the form of a trait or a product.

(x) Large-scale production of the desired product. 

5. Construction of an artificial recombinant DNA molecule has been made possible by the use of the following: 

(i) Plasmid is an independently replicating circular extra-chromosomal DNA. It contains only a few genes and it is generally found in bacterial cells. In nature, plasmids pass from one bacteria to another resulting in recombination.

(ii) Vector is an agent that carries a DNA fragment into Plasmids are used as vectors just like a mosquito acts as a host cell. insect vector to transfer the malarial parasite into the human body. 

(iii) Restriction enzymes are molecular scissors that cut a long DNA at specific points, into smaller fragments. In nature, bacterial cells use these enzymes to destroy the foreign DNA of viruses that attack them. 

(iv) DNA ligase is an enzyme that can rejoin cut fragments of DNA by sealing the sugar-phosphate backbone. Fragments of DNA can be from two different sources. 

(v) DNA polymerase is an enzyme that joins nucleotide monomers together to make a new DNA strand, using another strand as a template. 

6. The process of Construction of a Recombinant DNA 

(1) Suppose, a bacterium, Salmonella typhimurium, has a gene for antibiotic resistance. 

(ii) This antibiotic resistance gene is cut out of its DNA molecule with a restriction enzyme.

(iii) The cut piece of DNA is separated out (isolated).

(iv) It is then linked with a plasmid DNA using a DNA ligase enzyme.

(v) This makes a new combination of circular DNA created in vitro and is known as recombinant DNA. It is capable of autonomous replication inside a host cell.

The recombinant DNA if transferred into Escherichia coli, a bacterium closely related to Salmonella, replicates using the new host’s DNA polymerase enzyme and makes multiple copies or may even confer antibiotic resistance to the new host.

7 Tools of Recombinant DNA Technology The key tools required for recombinant technology are

(i) Restriction enzymes

(i) Ligases

(iii) Polymerase enzymes

(iv) Vectors

(v) Host organism/cell

8. Restriction Enzymes (REs) are normally the definite arsenal of bacteria against viral attack, as they cut their DNA into pieces thereby restricting the growth of invading bacteriophages.

(i) They cut the DNA at specific base sequences, known as recognition sequences. 

(ii) Biotechnologists use REs for cutting big chromosomal DNA into smaller fragments that can be used to study the role of genes or can be used to create recombinant DNA molecules.

(iii) The convention for naming these enzymes is 

(a) the first letter of the name comes from the genus.

(b) the second two letters come from the species of the prokaryotic cell from which they were isolated.

(iv) For example, in Eco RI, the letter ‘E’ is the first letter from Escherichia, ‘co’ are the second two letters of ‘coli’ and ‘R’ is derived from the name of the strain. 

Roman numbers following the names indicate the order in which the enzymes were isolated from that strain of bacteria. 

(vi) Restriction enzymes are of two kinds-exonucleases and endonucleases. (a) Exonucleases remove nucleotides from the outer ends of the DNA.

(b) Endonucleases make cuts at specific positions within the DNA. Each restriction endonuclease inspects the length of a DNA sequence. Once it finds its specific recognition sequence, it will bind to the DNA and cut each of the two strands of the double helix at specific points in their sugar-phosphate backbones.

(vii) Each restriction endonuclease recognises a specific palindromic nucleotide sequence (a sequence of base pairs that reads the same on the two strands when the orientation of the reading is kept the same). For example, the following sequence reads the same on the two strands whether read admin 5’3′ direction or 3′ 5′ direction.

(viii) Restriction enzymes cut the strand of DNA a little away from the centre of the palindrome sites, but between the same two bases on the opposite strands. These staggered cuts leave single-stranded portions at both ends. These are referred to as sticky ends.

(ix) Stickiness is the chemical ability to base pair with any other DNA molecule that has also been cut by the same RE. It means it will have the same sequence hanging unpaired. This stickiness of the ends facilitates the action of the enzyme DNA ligase. 

image 4

9. Use of Restriction Endonucleases

These enzymes are used in genetic engineering to cut the large DNA molecule into smaller fragments. When DNA from two different sources are cut by the same restriction enzyme, the resultant DNA fragments have the same kind of sticky ends and these can be joined together (end-to-end) using DNA ligases.

This new DNA created by joining fragments, from two different sources/genomes together, is recombinant DNA.

10. Separation of restriction fragments from each other can be done by gel electrophoresis.

(i) An electric field is applied and these fragments are forced through a viscous gel of agarose (a natural polymer extracted from seaweeds).

(ii) Since DNA fragments are negatively charged (because of their phosphate groups) they will move towards the positively charged pole. 

(iii) Smaller fragments move faster and bigger ones move slow, but they all separate out (according to their length) into bands that can be identified later by staining. 

(iv) The staining of DNA bands can be done with a compound known as ethidium bromide followed by exposure to UV radiation. Bright orange-coloured bands of DNA are visible.

A typical agarose gel electrophoresis showing migration of undigested (lane 1) and digested sets of DNA fragments (lanes 2 to 4) 

(v) The separated bands of DNA are cut out from the agarose gel and extracted from the gel piece. This step is known as the Solution.

The DNA fragments purified in this way are used in constructing recombinant DNA by joining them with DNA from plasmid vectors. 

11. Cloning Vectors Plasmids and bacteriophages have the ability to replicate within the bacterial cells independent of in chromosomal DNA, so they are used as cloning vectors. 

(i) They use the machinery of bacterial cells, to replicate and thereby increase the copy number (make clones) of the DNA inserted into them.

(ii) Purpose of cloning is to enhance gene expression and also to isolate these genes in large numbers to study their structure and function. 

(iii) Vectors are engineered in such way that they help easy linking of foreign DNA and allow selection of recombinants (bacterial cells that have picked up recombinant plasmid) only from non-recombinants (those who have not).

12. Features that are required to facilitate cloning into a vector 

(1) 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.

(ii) Selectable marker The selectable marker helps in identifying and eliminating non-transformants and selectively permitting the growth of the transformants. Transformation is a procedure through which a piece of DNA is introduced in a host bacterium. Normally, antibiotics such as ampicillin, chloramphenicol, tetracycline or kanamycin, etc., are considered useful selectable markers.

(iii) Cloning sites In order to link the alien DNA, the vector needs to have a cloning or recognition site for the commonly used restriction enzymes.

(a) For example, we can use E. coli cells that harbour plasmid with ampicillin and tetracycline resistance genes. (b) This E. coli cloning vector pBR322 is called a transformant as it harbours antibiotic resistance gene-carrying plasmid.

(c) The ligation of foreign DNA is carried out at a restriction site present in one of the two antibiotic-resistance genes. It can be done at the BamHI site of the tetracycline resistance gene. 

(d) In this case, the recombinant plasmids will lose tetracycline resistance due to the insertion of foreign DNA but can still be selected out from AMO non-recombinant ones by plating the transformants on a medium containing ampicillin.

(e) The recombinants will grow in an ampicillin-containing medium but not on that containing tetracycline. But, non-recombinants will grow on the medium containing both antibiotics.

(f) Other selectable markers can differentiate recombinants from non-recombinants on the basis of their ability to produce colour in the presence of a chromogenic substrate.

(G) The presence of a chromogenic substrate gives blue-coloured colonies if the plasmid in the bacteria does not have an insert.

(h) The tumour-inducing (Ti) plasmid of Agrobacterium tumefaciens has now been modified into a cloning vector that is able to use the mechanisms to deliver genes of our interest into a variety of plants.

(I) Retroviruses can be disarmed and used to deliver desirable genes into animal cells.

13. Since DNA is a hydrophilic molecule, it cannot pass through a cell membrane that has hydrophobic ends on both inside and outside. Hence, various artificial means used to make the cells take up foreign DNA are 

(i) DNA is coated with lipids. Fusion of this lipid and the membrane lipid can occur, facilitating the entry of DNA. 

(ii) Sometimes, bacteria are treated with a specific concentration of a divalent cation, such as calcium, which makes the cell membrane more permeable. Recombinant DNA can then be forced into such cells by incubating the cells with recombinant DNA on ice, followed by placing them Briefly at 42°C (heat shock) and then putting them back on ice. This enables the bacteria to take up the recombinant DNA. 

(iii) Recombinant DNA is directly injected into the nucleus of an animal cell, using a microsyringe. This is known as micro-injection. 

(iv) Plants’ cells are bombarded with high-velocity micro-particles of gold or tungsten coated with DNA in a method known as biolistics or gene gun.

(v) Some pathogens that naturally infect a cell can be ‘disarmed’ (by eliminating their harmful gene) and then allowed to infect the cell, carrying the desired recombinant DNA into the host. 

(vi) In eukaryotic cell, since the term transformation is used To hold for cancerous cells, therefore introduction of foreign DNA to a eukaryotic cell is called transfection.

14. Processes of Recombinant DNA Technology lim As has been listed already, the recombinant DNA technology involves 

(i) isolation of DNA.

(ii) fragmentation of DNA by restriction of endonucleases. 

(iii) isolation of a desired fragment of DNA.

(iv) ligation of the DNA fragment into a vector.

(v) transferring the recombinant DNA into the host. 

(vi) culturing the host cells in a medium at a large scale. 

(vii) extraction of the desired product.

15. Isolation of Genetic Material (DNA) The genetic material, which is deoxyribonucleic acid or DNA needs to be in pure form, free from other macromolecules, and able to act on it. The following are the steps: 

(i) Cell wall needs to be broken to release DNA. For this, the bacterial cells/plant or animal tissue are treated with enzymes such as lysozyme (bacteria), cellulase (plant cells), and chitinase (fungus). REs to be

(ii) Genes are located on long molecules of DNA tightly linked with histone proteins.

(iii) Histones and other proteins have to be removed by treatment with proteases.

Since, other macromolecules such as RNA, proteins, polysaccharides and lipids, also come out along with DNA, they have to be removed by appropriate treatments.

(iv) With the addition of chilled alcohol, purified DNA precipitates out.

16. Cutting of DNA at Specific Locations 

(1) Enzyme cuts the purified DNA into fragments.

(ii) Progression of restriction enzyme digestion is monitored through agarose gel electrophoresis. 

(iii) The cutout gene of interest from the source DNA and the vector with a cut space are mixed and ligase is added. The sticky ends of cut DNA from both sources binds

hydrogen bond with each other and ligase seals the nicks. The resulting recombined molecule is called recombinant DNA.

17. Amplification of Gene of Interest using PCR PCR stands for Polymerase Chain Reaction, a method of amplifying fragments of DNA. This method can make multiple copies of even a single DNA fragment or the gene interest, in a test tube.

The reaction mixture requires 

(1) Double-stranded DNA fragment (gene of interest).

(ii) Primers are small chemically synthesised oligonucleotides that are complementary to the regions of this DNA. 

(iii) The special thermostable DNA polymerase (isolated from bacteria, Thermus aquaticus), that does not denature and remains active even at high temperatures.

18. During each cycle of PCR, the following occurs: 

(i) Denaturation of DNA sample Unwinding of two strands of DNA by heating the sample at 92-94°C. 

(ii) Primer annealing Primers get positioned on the exposed nucleotides as per base pairing rules. 

(ii) Extension of primers DNA polymerase recognises primers as ‘start’ tags and begins to extend the primers using the free nucleotides provided in the reaction and the genomic DNA as a template,quar

(iv) With each round of reactions, the DNA doubles. If the process of replication of DNA is repeated many times, the segment of DNA can be amplified to approximately a billion times, i.e., 1 billion copies are made, Jamal

(v) The amplified fragment if desired can now be used to ligate with a vector for further cloning

19. Insertion of Recombinant DNA into the Host 

(1) If a recombinant DNA bearing gene for the resistance to Cell/Organism an antibiotic (e.g., ampicillin) is transferred into E. coli cells, the host cells become transformed into ampicillin-resistant cells. 

(ii) If we spread the transformed cells on agar plates containing ampicillin, only transformants will grow, and untransformed recipient cells will die. 

(iii) Due to the ampicillin resistance gene, it is possible to select a transformed cell in the presence of ampicillin. The ampicillin resistance gene in this case is a selectable marker. s called a

20. Obtaining the Foreign Gene Product Fam

(1) The foreign gene gets expressed under appropriate conditions. After having the cloned gene of interest and having optimised conditions to induce the expression of the target protein, large-scale production can be carried out. 

(ii) If any protein-encoding gene is expressed in a heterologous host, the product protein is called a recombinant protein.

(iii) The cells harbouring cloned genes of interest may be grown on a small scale in the laboratory. The cultures may be used for extracting the desired protein and then

purifying it by using different separation techniques.

(iv) The cells can also be multiplied in a continuous culture system, wherein the used medium is drained out from one side while the fresh medium is added from the other to maintain the cells in their physiologically most active log/exponential phase.

(v) This type of culturing method produces a larger biomass leading to higher yields of the desired protein. 

(vi) For large-scale production, bioreactors are used of culture can be processed. Bioreactors are large vessels (100-1000 L capacity) in which raw materials are biologically converted into specific products, individual enzymes, etc., using microbial, plant, animal or human cells.

(vii) A bioreactor is specially designed to provide the optimal conditions of temperature, pH, substrate, salts, vitamins, oxygen, etc., for achieving the desired production levels. 21. Downstream Processing

After completion of the biosynthetic phase, the product is subjected to a series of processes to make it a market-ready product. These processes, collectively referred to as downstream processing, include

(1) Separation and purification.

(ii) Formulation with suitable preservatives.

(iii) Clinical trials as in the case of drugs. 

(iv) Quality control testing, etc.


biotechnology principles and processes neet questions


Question 1. Can you list 10 recombinant proteins which are used in medical practice? Find out where they are used as therapeutics (use the internet).

Answer

image

Question 2. Make a chart (with diagrammatic representation) showing a restriction enzyme, the substrate DNA on which it acts, the site at which it cuts DNA and the product it produces.

Question 3. From what you have learnt, can you tell whether enzymes are bigger or DNA is bigger in molecular size? How did you know?

Answer DNA is bigger in molecular size than enzymes. Because DNA is a long double-stranded molecule that can go up to a few meters in length when stretched end to end but enzymes although variable in size, would still be smaller than the DNA.

Question 4. What would be the molar concentration of human DNA in a human cell? Consult your teacher

Answer The molar concentration of human DNA in a human cell is 3.2 picograms. 

Question 5. Do eukaryotic cells have restriction endonucleases? Justify your answer

Answer No, eukaryotic cells do not have restriction endonucleases. This is because the restriction endonucleases are a defence mechanism to destroy foreign DNA. If it invades the cell, it is generally prokaryotes that get infected by naked viral and bacterial DNA that needs to be destroyed.

Question 6. Besides better aeration and mixing properties, what other advantages do stirred tank bioreactors have over shake flasks? 

Answer Besides better aeration and mixing properties, stirred tank bioreactors have many features that help maintain optimal conditions of temperature, pH, substrate, salts, vitamins, oxygen, etc., and make them more advantageous than a shake flask. These features are

(i) agitator system.

(ii) oxygen delivery system. 

(iii) foam control system.

(iv) temperature control system. 

(v) pH control system.

(vi) sampling ports so that small volumes of the culture can be withdrawn periodically.

Question 7. Collect five examples of palindromic DNA sequences by

consulting your teacher. Better try to create a palindromic sequence by following base-pair rules.

Answer

(1) 5-AAGCTT-3′ 3′-TTCGAA-5

(ii) 5′-GAATTC-3 3-GAATTC-5′

(i) 5′-ACTAGT-3′

3-CTTAAG-57

(iv) 5′-ACGCGT-3′ 3-TGATCA-30

(v) 5′-CCTAG G-3′ 3′-GGATCC-5′

Question 8. Can you recall meiosis and indicate at what stage a recombinant DNA is made?

Answer: Recombinant DNA is made at the pachytene stage during the end of prophase-1 when homologous chromosomes come together after duplicating and lie as packs of four.

They are physically close to cross over and exchange parts of their chromosome. This results in the recombination of DNA in homologous chromosomes.

Question 9. Can you think and answer how a reporter enzyme can be used to monitor the transformation of host cells by foreign DNA in addition to a selectable marker?

Answer: A reporter enzyme can be used to monitor the transformation of host cells by keeping track of its receptor gene. Examples of reporter enzymes are B-galactosidase or alkaline phosphatase. They have particular characteristics that allow visual or spectrophotometric detection of the activity of their corresponding genes. 

If host cells are transformed by the uptake of foreign DNA, that will cause insertional inactivation of the enzyme gene, the gene product will not express and colonies of host cells will appear colourless in the presence of a chromogenic substrate, whereas non-transformed cells will make blue coloured colonies as their enzyme will be active,

Question 10. Describe briefly the following 

(a) Origin of replication

(b) Bioreactors

(c) Downstream processing

Answer

(a) Origin of replication (ori) is a sequence on the chromosome, 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 also controls the copy number of the linked DNA. So, if we want to recover many copies of the target DNA it should be linked to the ‘ori’ site and should be cloned in a vector whose origin supports high copy number.

(b) Bioreactors are large vessels in which raw materials are biologically converted into specific products, individual enzymes, etc., using microbial, plant, animal or human cells.

A bioreactor provides the optimal conditions for achieving the desired production levels by providing optimum growth conditions of temperature, pH, substrate, salts, vitamins, oxygen, etc.

(c) Downstream processes include separation and purification, formulation with suitable preservatives, etc., which are collectively referred to as downstream processing. Such formulation has to undergo clinical trials as in the case of drugs,

Strict quality control testing for each product is also required. The downstream processing and quality control testing vary from product to product.

Question 11. Explain briefly

(a) PCR

(b) Restriction enzymes and DNA 

(c) Chitinase

Answer

(a) PCR stands for Polymerase Chain Reaction, a method of amplifying fragments of DNA. This method can make multiple copies of even a single DNA fragment or the gene of interest, in a test tube. The reaction mixture requires

(1) Double-stranded DNA fragment (gene of interest).

(ii) Primers-small chemically synthesised oligonucleotides that are complementary to the regions of this DNA. 

(iii) The special thermostable DNA polymerase (isolated from a bacterium, Thermus aquaticus), that does not denature and remains active even at high temperatures.

Unwinding two strands of DNA by heating the sample at 92-94°C helps primers to get positioned on the exposed nucleotides as per base pairing rules. DNA polymerase recognises primers as ‘start’ tags and begins to extend the primers using the free nucleotides provided in the reaction and the genomic DNA as a template. With each round of reactions, the DNA doubles.

(b). These enzymes are used in genetic engineering to cut the large DNA molecule into smaller fragments.

When DNA from two different sources are cut by the same restriction enzyme, the resultant DNA fragments have the same kind of atbalsticky-ends’ and these can be joined together (end-to-end) using DNA ligases.

This new DNA is created by joining fragments, from two different sources/genomes together is recombinant DNA. 

(c) Chitinase is an enzyme that breaks down chitin, a component of fungal cell wall, It is useful for isolating the fungal cell DNA.

Question 12. Discuss with your teacher and find out how to distinguish between

(a) Plasmid DNA and chromosomal DNA?

(b) RNA and DNA?

(c) Exonuclease and endonuclease?

Answer

(a) Plasmid DNA and chromosomal DNA

image 1
credit- Brainly.in

(b) RNA and DNA

image 2 1024x712 1

(c) Exonuclease and endonuclease

image 3
credit- Brainly.in

biotechnology principles and processes ncert solutions 


Question 1. How is the copy number of the plasmid vector related to the yield of the recombinant protein? 

Ans:- The recombinant DNA can multiply as many times as the copy number of the plasmid vector thereby determining the yield of recombinant protein. So, higher the copy number of the plasmid vector, higher the yield recombinant protein.

Question 2. What does ‘H’ in ‘d’ and ‘III’ refer to in the enzyme HindIII?

Answer

(i) The first letter ‘H’ indicates the genus of the organism from which the enzyme was isolated H = genus Haemophilus. (ii) The fourth letter ‘d’ indicates the particular strain used to produce the enzyme-d = strain Rd.

(ii) The Roman numerals denote the sequence in which the restriction endonuclease enzyme from that particular genus, species and strain of bacteria have been isolated-III, i.e., third restriction endonuclease to be isolated from this species.

Question 3. Do biomolecules (DNA and protein) exhibit biological activity in anhydrous conditions?

Answer No, for DNA to function, proteins in the form of enzymes are required which do not work in anhydrous conditions. That is why no life activity can take place in anhydrous conditions.

Question 4. What modification is done on the Ti plasmid of Agrobacterium tumefaciens to convert it into a cloning vector? 

Answer Tumour inducing (Ti) plasmid of Agrobacterium tumefaciens is modified into a cloning vector by removing/altering the gene responsible for its pathogenic property to the plants. This is so that it does not cause harm but can act as a delivery system for genes of interest into a variety of plants and other cells.


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