Tians ENGINEERING | MEDICAL| FOUNDATION RANKERS ™ = NEET BIOLOGY bey) JEE MAIN Bi) JEE ADVANCED EZ7Z4NEET UG JEE Main/Advanced, NEET, BITSAT, ISAT, VITEEE, EAMCET, UPTU SEE, Orissa JEE, WBJEE, SRMEEE, MHT CET, Delhi CEE, CUSAT CAT, AMU, KEAM, VTU, TANCET, ENAT, COMEDK UGET, PTU CETFROM THE DESK OF DIRECTORS ADITYA SINGHAL Promoter Director asklITians Web Pvt. Ltd. NISHANT SINHA Promoter Director askliTians Web Pvt. Ltd. Every child is future of the family and the nation. In today’s world of cut-throat competition and minute specialization, the toughest task parents and teachers face is to mold our children to face the challenges and uncertainties that lie ahead Letus stand with them ensuring their best throughout the school examinations and for the competitive exams. Let us be highly focused and concemed about broader educational objectives of developing _ students’ confidence, self motivation and thinking capability to benefit their future Inspiration is what askilTians has been giving to students throughout its wonderful journey of 13 Years. Be it an IIT aspirant or students with aspirations for medical, it's the inspiration of interacting and taking - guidance from their role models is what drives our student's motivation. askllTians believes that students already have the Necessary potential in them, and as a career shaping Partner of the student, it never failed to take out the best Potential out of them.Classroem Asus | “AM Package BIOLOGY Module - 10 S.No. / Chapter Name Page No. Biotechnology : Principles and Processes 1029-1049 Biotechnology and its Applications 1050-1066Number of Questions Trend Analysis AIPMT/ NEET 2010 2011 2012 2013 2014 2015 2016 2017 2018 2019 Year Chapter Utility Score 45/5 87(103g INTRODUCTION Biotechnology may be defined as use of micro-organisms animals, or plant cells or their products to generate different products at industrial scale and services useful to human beings. Old biotechnology are based on the natural capabilities of ‘micro-organisms. 8g, formation of citrie acid, production of penicillin by Penicittium notatu. New biotechnology is based on recombinant DNA technology. E.g., Human gene producing insulin has been transferred and expressed in bacteria like E.coli In modern biotechnology, different types of valuable products are produced with help of microbiology, biochemistry, tissue culture, chemical engineering and ‘genetic engineering, molecular biology and immunology. GENETIC ENGINEERING Genetic engineering (also referred as recombinant DNA technology or gene splicing) is one kind of biotechnology involving manipulation of DNA. It deals with the sol of useful genes from a variety of sources and the formation ‘fnew combinations of DNA (recombinant DNA) for repair, improvement, perfection and matching ofa genotype. Genetic engineering may be defined as a technique for artificial and deliberately modifying DNA (gene) to suit human needs, In genetic engineering, for manipulation, DNA molecule in out at two desired places with the help of restriction ‘endonuclease to isolate a specific DNA segment and then inserted in another DNA molecule at a desired position, ‘The new DNA molecule is recombinant DNA and the technique is called genetic engineering. Genetic engineering aims at adding, removing or repairing ofa part of genetic material Paul Berg (Father of genctic engineering) transferred gene of SV40 virus (simian virus) into E.coli with the help of 2. phage (Nobel prize - 1980). ‘The concept of genetic engineering was the outcome of ‘wo very significant discoveries made in bacterial research. ‘These were — Presence of extrachromosomal DNA fragments called plasmids inthe bacterial cell, which replicate along with ‘chromosomal DNA ofthe bacterium, — Presence of enzyme restriction endonucleases which ‘cut DNA at specifi sites. These enzymes are, therefore, called molecular scissors. zee} TOOLS AND TECHNIQUES OF RECOMBINANT DNA TECHNOLOGY ENZYMES ‘A number of specific kinds of enzymes are employed in genetic engineering, Lysing enzymes : These enzymes are used for opening the cells to get DNA for genetic experiment. Bacterial cell wall is commonly dissolved with the help of lysozyme, Cleaving enzymes : These enzymes are of two types- cexonucleases and endonucleases. ~ Exonucleases cut off nucleotides from 5’ or 3' end of DNA molecule. ~ _ Endonucteases break DNA duplex at any point except the end. ~ Restriction endonucleases are endonucleases that cleave DNA duplex at specific points in such a way that they come to possess short single-stranded free ends. ~ Restriction enzyme (EcoRI) was discovered by Arber, ‘Smith and Nathans (1978 Nobel prize). These enzymes exist in many bacteria. Besides cleavage, some restriction endonuclease also have the capability of ‘modification. Restriction enzymes are used in recombinant DNA. technology because they can be used in vitro to recognise and cleave within specific DNA sequence ‘typically consisting of 4 to8 nucleotides. This specific 4 108 nucleotide sequence is called restriction siteand is usually palindromic, this means that the DNA. ‘sequence isthe same when read in a5'->3' direction on both DNA strands. AND MADAM DNA Asa result, the DNA fragments produced by cleavage with these enzymes have short single-stranded overhang at each end, These kinds of ends are called sticky or cohesive ends because base pairing between them can stick the DNA molecules together again, 4 GAATTC3' CTTAAGS) ¥ 4 S'AATTCS 365 DTTAAS! Therefore, by cutting two different DNA samples with the same restriction enzyme and mixing the fragments together, a recombinant DNA molecule can be ‘generated.Exceptionally, some enzymes cleave both strands of DNA exactly atthe same nucleotide position, typically inthe centre ofthe recognition sequence resulting in blunt end or fush end, Sma I (Serratia marcescens) ¥ sccCcGGGy scccs+ 6663 seaeeces 3GGGS+3'CCC3 oN «= Learn More a Isoschizomers Isoschizomers are restriction endonucleases that recognise the same DNA sequence and makerthe same cut. Hpall (recognition sequence: CCGG) and Msp (recognition sequence: C?CGG) areisoschizomers, ~The convention for naming restriction enzymes is the first letter is from the genus and the next two letters are from the species of prokaryotic organism followed the name of the strain and roman number indicating the order of isolation of the enzyme. — These fragments can be separated by a technique known as gel electrophoresis, a method that exploits the fact that these molecules carry charged groups that cause them to migrate under an electric field through a matrix. DNA fragments separate according to their size, with smaller DNA fragments further from the end ofthe gel where sample was loaded, ~ Themost commonly used matrix is agarose, a natural polymer extracted from sea weeds. The separated DNA fragments can be visualised as bright orange coloured bands only after staining the gel with a compound known as ethidium bromide followed by exposure to UV radiations. = Theseparated bands of DNA are cut out from agarose gol and extracted from the gel piece, This tep is called elution, wells DNA a ny y_Lanest Smallest oO fii Ost lt] WHE oO Fig. : Atypical agarose gel electrophoresis showing migration ‘ofundigested (lane 1) and digested set of DNA fragments (lane2to4) —_ Synthesising enzymes : These enzymes are used to synthesise new strands of DNA, complementary to existing DNA or RNA template. They are of two types: reverse transcriptases and DNA polymerases. — Reverse transcriptases help in the synthesis of complementary DNA strands on RNA templates. — DNA polymerases help in the synthesis of complementary DNA strands on DNA templates. Action of Restriction enzyme The enzyme cuts both DNA Strands al the same site Vector DNA, ROSS: BeoRI ‘Sticky end Bok euts the Gand A only when the sequence INA between bases GAATTC is present sn the DNA, Foreign DNA EH OWT Sticky end DNA fragments join at sticky ends RII PIV Recombinant DNA ‘Steps in the formation of recombin nt DNA by ction of restriction endonuclease enzyme - EcoRI— Joining enzymes : These enzymes help in joining the DNA fragments. For example, DNA ligase from Escherichia coli is used to join DNA fragments Joining enzymes are, therefore, called molecular glues. eaeeas CLONING VECTORS Alkaline phosphatases : These enzymes cut off + phosphate group from the 5'end of linearised circular DNA and prevent its recircularisation. seam or Southern Blotting Principle — Southern blotting is a hybridisation technique for identification of particular sizeof DNA from the mixture ‘of other similar molecules. It was developed by Edward M. Southern (1975). This technique is based on the principle of separation of DNA fragments by gel electrophoresis and identified by labelled probe hybridisation — Basically, the DNA fragments are separated on the basis of size and charge during electrophoresis. Separated DNA fragments are transferred on nylon membrane and then the desired DNA is detected using specific DNA probe that is complementary othe desired DNA. — A hhybridisation or DNA probe is short (100-S00bp), single-stranded DNA. The probes are labeled with a ‘marker so that they can be detected after hybridisation. Procedure ~ Restriction digest: bly RE enzyme and amplification by PCR — Gelelectrophoresis: SDS gel electrophoresis ~ Denaturation: Treating with HCl and NaOH — Blotting: Transferring digested DNA to nylon membrane ~ Baking and Blocking with casein in BSA. — Hybridisation using labelled probes — Visualisation by autoradiogram Application ~ Used to detect DNA in given sample ~ Used for paternity testing, criminal identification, victim ‘identification ~ Toisolate and identify desire gene of interest ~ Used in restriction fragment length polymorphism (RFLP) — To identify mutation or gene rearrangement in the sequence of DNA — Used in diagnosis of disease caused by genetic defects ~ Used to identify infectious agents ‘The DNA used as a carrier for transferring a fragment of foreign DNA into. suitable host is called vehicle or vector DNA. Plasmids and bacteriophages have the ability to replicate within bacterial cells independent of the control of chromosomal DNA. ‘The followingare the features that are required to facilitate cloning into a vector: = Origin of replication (ori) : This is a sequence from ‘where replication starts. The vector requires, an origin ‘of replication (or so that it is able to multiply within the host cells. This sequence is also responsible for ‘controlling the copy number of the inked DNA. This implies that any foreign DNA inserted into this vector will also be replicated in the process Cla Hind 1 oli cloning vector pBR322 showing restriction sites, ori and antibiotic resistance ~ Selectable marker : Along withor' the vector requires a selectable marker for identification of transformed host cells from non-transformed cell. Normally, the genes encoding resistance to antibioties such as ampicili, chloramphenicol, tetracycline orkanamyein, etc., are considered useful selectable markers for E. coli. ~ Cloning sites: In order tolinkthealien DNA, the vector needs recognition sites for the commonly used restriction enzymes. The ligation of alien DNA is carried out at a restriction site present in one of the two antibiotic resistance genes. For example, if foreign DNA is lighted at the Bam HI site of tetracycline resistance gene inthe vector pBR322, the recombinant plasmids loss tetracycline resistance due to insertion of foreign DNA (insertional inactivation). Now, itean be selected out fom non-recombinant ones by plating the transformants on ampicillin-containing medium. The‘transformants (plasmid transfer) growing on ampicillin- containing medium are then transferred on a medium containing tetracycline. The recombinants will grow in ampicillin-containing medium but not on that containing tetracycline. But, non-recombinants will ‘grow on the medium containing both the antibiotics. In this ease, one antibiotic resistance gene helps in selecting the transformants. Due to inactivation of antibiotic resistance selection ‘ofrecombinants is a troublesome 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, recombinant DNA is inserted within the coding sequence of an enzyme, leading to its inactivation which is referred to as insertional inactivation. The presence ofa chromogenic substrate X-gal (5-bromo-4-chloro-3-indohyl B-D galactopyranoside) gives blue-coloured colonies ifthe plasmid inthe bacteria does not have an insert, Presence ‘of insert results into insertional inactivation of the - galactosidase (reporter enzyme) and the colonies do not produce any colour, these are identified as recombinant colonies. — Vectors for cloning genes in plants and animal : Agrobacterium tumefaciens delivers a piece of DNA (known as T-DNA) to transform normal plant cells into a tumour, Similarly, retroviruses in animals have the ability to transform normal cells into cancerous cells. So, modified Ti plasmid from Agrobacterium and disarmed retrovirus have been used as cloning vectors to deliver gene of interest into plant and animal cells, respectively. Types of Cloning Vectors Plasmids ‘These are extrachromosomal DNA segments found in bacteria which can replicate independently. Plasmids can be taken out of bacteria and made to combine with desired DNA segments by means of restriction enzymes and DNA ligase. A plasmid carrying DNA of another organism integrated with it isknown as recombinant plasmid or hybrid plasmid or chimeric plasmid, Eg. pBR vector plasmids (named after the discoverer Bolivar ‘and Rodriguez, pUC vector plasmid University of California). Viruses ‘The DNA of certain viruses is also suitable for use as a vehicle DNA. Bacteriophage (bacterial virus) has been used to transfer gene for B Galactosidase from Escherichia col to human calls. [Lambda phage (2 phage) has been used for transferring lac genes of E.coli into haploid callus of tomato, ‘Table: Some vectors and their insert: eaaceie Insert size kb Plasmid 05-8 Bacteriophage lamda 923 Cosmid 30-45 BAC 50-300 vac 1000-2500 Antibiotic resistance gene Centromere Origin of replication Telomere (Ori) Plasmi Yeast Artificial ‘Chromosome (YAC) TARGET DNA Itisthe DNA which is transferred from one organism into another by combining it withthe vehicle DNA. The passenger DNA can be complementary, synthetic or random. ‘Complementary DNA (¢DNA) :Itis synthesised on mRNA, template withthe help ofreverse transcriptase and necessary nucleotides. Synthetic DNA (sDNA) : It issynthesised with the help of DNA polymerase on DNA template. Random DNA : It refers to small fragments formed by breaking # chromosome with the help of restriction ‘endonucleases. COMPENENT HOST ‘Transformation isa process through which a DNA fragment is introduced in a host cell. For uptake & hydrophilic DNA, bacterial host cell are treated with specific concentration of a divalent cation such as calcium to make pores in the cell wall. This is followed by incubation of competent cell with. recombinant DNA on ice followed by placing them briefly at 42°C (heat-shock) and then putting them back on ice. Microinjection method is another way to transfer ‘recombinant DNA into host cells In this method, recombinant DNA is directly injected into the nucleus of an animal cell. In plants, biolistics or gene gun is used. In this method, cells are bombarded with high-velocity micro-particles of gold or ‘tungsten coated with DNA.£ s ie representation of recombinant DNA technology Fi : Diagramma PROCESS OF RECOMBINANT DNA TECHNOLOGY ‘A recombinant DNA molecule is produced by joining together of two or more DNA segments usually originating from different organisms. More specifically a recombinant DNA molecules a vector (e., plasmid, phage or virus) into which the desired DNA fragment has been inserted to enable its cloning in an appropriate host. Recombinant DNA molecules are produced with one ofthe following three objectives = To obtain a large number of copies of specific DNA fragments = Torecover large quantities ofthe protein produced by the concemed gene. ~ Tointegratethe gene in question into thechromosome ‘ofa target organism where it expresses itself. ‘This technique developed by genetic engineering. In this technique, frst of all isolation of desired gene from any organism is carried out followed by its transfer and «expression into any organism of choice, They are known as ‘transgenie micro-organisms, Transgenic miero-orzanisms are produced with view to obtain novel pharmaceutical proteins. Forexample- Human insutn isbeing produced commercially from transgenic E.coli strain, Many valuable recombinant proteins ae also being produced using transgenic animal cll lines and transgenic plants. [At the same time, a number of these proteins of great ‘medicinal value could not be produced on a commercial scale using the non-transgenic cells or organisms. Proteins produced by transgenes are called recombinant proteins, Such type of recombinant genes are utilised for the formation of different products. ‘+ Application ofrecombinant DNA technology : The technique ‘of recombinant DNA can be employed in the following ways: It can be used to elucidate molecular events in the biological process such as cellular differentiation and ageing. The same can be used for making gene maps ‘with precision, — In biochemical and pharmaceutical industry, by engineering genes, useful chemical compounds ean be produced cheaply and efficiently. — Production of transgenic plants, — Production of genetically modified micro-organisms, STEPS OF RECOMBINANT DNA TECHNOLOGY Recombinant DNA technology involves several steps in specie sequence such as Isolation ofa genetic material = Cutting of DNA at specific locations — Amplification of gene of interest using polymerase chain reaction = Insertion of recombinant DNA into the host cell/ organism. ~ Obtaining the foreign gene product — Downstream processingjiotechnology: Principles and Processes Isolation of Genetic Material (DNA) + Theremoval ofplasmid or genomic DNA fiom cellsis termed isolation. + Isolation usually involves the breaking of the cell’s ‘membrane (and possibly nuclear membrane) and possibly also cell wall (plant cells), DNA is released from the cell by ‘treating the bacterial cells/plant or animal tissue with enzymes such as lysozyme (bacteria), cellulase (plant cells), chitinase (fungus). Then, RNA is removed by treating with ribonuclease while proteins is removed by treating with protease. + Other molecules are removed by suitable treatments. The purified DNA ultimately precipates out as collection of fine threads after the addition of chilled ethanol. a ih Fig. : DNA that separates out can be removed by spooling Cutting of DNA at Specific Locations + DNA purified from an organism can be prepared for cloning only after it has been cut into smaller molecules. + Restriction endonucleases make the cleavage of DNA possible. + Agarose gel electrophoresis is employed to check the progression ofa restriction enzyme digestion. The process is repeated with the vector DNA also. After having cut the source DNA as well as the vector DNA with a specific restriction enzyme, the cut out 'gene of interest’ from the source DNA and the cut vector with space are mixed and. ligase is added. DNA ligase joins DNA to DNA. This results in the formation of recombinant DNA. Amplification of Gene of Interest Using Polymerase Chain Reaction (PCR) + The polymerase chain reaction is a repetitive bidirectional synthesis of DNA. In this reaction, multiple copies of the gene (or DNA) of interest are synthesised in vitro using {wo sets of small chemically synthesised oligonucleotides called primers and the enzyme, DNA polymerase. + The thermostable DNA polymerase, Tag, DNA polymerase (Gsolated from Thermus aquaticus) extends the primers using, the nucleotides provided in the reaction and the genomic DNAs template. + Asamplification proceeds, the DNA sequence between the primers double after each cycle. The process of replication ‘of DNA is repeated many times. The segment of DNA can be amplified to approximately billion times, The amplified fragment can now be used to ligate with a vector for further cloning. oC tii. 0, SSeS. AV eons i” | 8 a conn EEE — ages Fig. Polymerase chain reaction (PCR) : Each cyele has three ‘steps: (2) Denaturation; (i) Primer annealing: and (ii) Extension of primers oD Ss a a Learn More = ‘Thermal eyeler “The thermal cycler (also known as a thermocycler, PCR Machine or DNA Amplifier) is a laboratory apparatus used to amplify ‘segments of DNA via the Polymerase Chain Reaction (PCR). ‘The device has a thermal block with holes where tubes holding the PCR reaction mixtures can be inserted. Insertion of Recombinant DNA into the Host Cell/ Organism + The desired DNA sequence, once attached to a DNA vector, must be transferred (o a suitable host. Transformation is defined as the introduction of foreign DNA intoa recipient cell. Transformation ofa cell with DNA froma virus isustally referred to as transfection. There are several methods of introducing the ligated DNA into reeipient cells. Escherichia col is usually the host, and transformation of E.coli is an ‘essential step in these experiments. Several methods are available for the transfer of DNA into cells of higher agian >’ Learn More Gene cloning ‘Gene cloning is the process in which a gene of interest is located and copied (cloned) out of DNA isolated from an organism, The DNA isolated from an organism contains thousands of different ‘genes. To find one specific gene coding the specifie protein of interest, scientists make gene libraries to catalogue the organism's DNA. The gene the scientist is looking for is selected from this library.ing the Foreign Gene Product ‘The ultimate aim of recombinant DNA technology is to Produce a desirable protein, Therefore, there is anced for the recombinant DNA to be expressed, The foreign gene gets expressed under appropriate conditions. The cultures may be used for extracting the desired protein and then purifying it by using different separation techniques. To produce desired proteins in large quantities, the development of bioreactors where large volumes (100-1000 litres) of culture can be processed, was required ‘Thus, bioreactors can be thought of as vessels in which raw materials are biologically converted into specific products, individual enzymes, ec., using microbial plant, animal or human cells. A bioreactor provides the optimal conditions for achieving the desired product by providing ‘optimum growth conditions (temperature, pH, substrate, salts, vitamins, oxygen). A.stirred-tank reactor is usually cylindrical or witha curved base to facilitate the mixing of the reactor contents, The stirrer facilitates even mixing and oxygen availability throughout the bioreactor. Alternatively, air can be bubbled through the reactor. The bioreactor has an agitator system, an oxygen delivery system and a foam control system, a ‘The most commonly used bioreactors are of stirring type. Motor Acidibase forpl contol Feed Iaker Flat bladed Sea for impeller Culture roth Sterile air {) Simple stirred-tank bioreactor eee} temperature contol system, pH controls and sampling arts othat smal volumes of th eulturcan be withdrawn periodically Micro-organisms can be grown in bioreactors in two ways: Support growth system : In tis method, micro- organisms are grown asa thin lye or fli he solid medio, Suspended growth system : By suspending els o myeala inthe liguid medium scaled suspend growth yan Manufacturing uni : During the designing of bioeactor forthe process, often very large sized units sed so that it accommodsie huge ameunt ofmediam, DOWNSTREAM PROCESSING + Afier completion of the biosynthetic stage, the product is subjected through a series of processes before it is ready for marketing as a finished product. The processes include separation and purification, which are collectively referred to as downstream processing. The product has to be formulated with suitable preservatives. This is followed by ‘thorough clinical trials (in case of drugs) are strict quality control testing, Bubbles dramatically inerese the oxygen ‘wansfer aes ) (b) Sparged stirred-tank bioreactor@ EXERCISE-1| TOPICWISE MCQs Restriction endonucleases (@) are enzymes that process pre-RNA. (©) areenzymes that degrade DNA. (©) protect bacterial cells fom viral infection. (@ None of the above ‘The primary reason why the same basic techniques can be used to analyse the DNA from species s diverse as bacteria. and bumans is that nif (@) allcels are identical Critical Thinkng (0) every organism has the same amount of DNA. (the DNA sequences of all organisms are the same. (®_DNAbas aconsistent structure in al organisms. 3. Which of the following isa critically important tool used in experiments involving DNA hybridisation? (@) DNA sequencing machines () Ligases (© DNAprobes (@) Vectors 4. A.gene is said tobe cloned if (@ the DNA sequence ofthe gene is known () the function ofthe gene is known. (©) thereis a DNA probe forthe gene. (@) the gene has been isolated and copied. 5. An enzyme that joins the ends of two strands of nucleic acids: (@ Polymerase (b) Synthetase (©) Helicase (@) Ligase 6. Introduction of one or more genes into an organism which normally doesnot possess them or their deletion by using artificial means (notbybreeding) comes under: Critical Thinking (@) Molecular biology (b) Cytogenetics ©. Genetichybridisaion @ Geneticengineering 7. AeDNAclone contains (@) mostly cytosine (©) copy of the DNA identical to the nuclear gene. (© copy of non-coding DNA (0 aDNA copy of mRNA. 8 Agenetiemarkerisa {@ place where a restrition enzyme cuts DNA. (b) chart that traces the family history ofa genetic trait (©) muceotide sequence near a particular gene (@) radioactive probe used to find a gene 2 13. 14, 1s. 16. 1 18. DNA fragments are separated using gel electrophoresis @) © © @ because DNA is pulled through the gel toward the negative end of the field because larger DNA fragments move faster through. the gel than smaller DNA fragments. toidentify and isolate DNA fragments. to synthesise DNA for cloning. First restriction endonuclease enzyme was discovered in: @) (b) © @ PPLO E.coli Hacmophilus influencae Bacteriophages: TThe enzyme which helps to cut one strand of DNA duplex torelease tension of coiling of two strands is: @ © Critical hg DNA ligase (6) DNA polymerase ‘Topoisomerase (@) Helicase or unwindase ‘Annucleie acid probe might be used to @ © © @ insert genes into a host cell. make DNA for gene cloning. splice pieces of DNA. find a nucleotide sequence. ‘The linking of antibiotic resistance gene with the plasmid vector became possible with: @ © (©) Endonucleases (@) Exonucleases DNA ligase DNA polymerase Reporter genes include genes for: (@) drug resistance (b) bioluminescence (© DNAorigins (@ Both (a) and (b) DNA fragments with sticky ends are not allowed to undergo self ligation by: (@) Unwindase () Singlestrand binding proteins (©) Gyrase (@)_ Alkaline phosphatase ‘Which of the following is corectly matched? @ ©) © @ Ligase— Molecular glue ‘Thermus aquaticus ~ Bt gene Agrobacterium tumefaciens — Cosmid veetor Hind I -Plasmitd vector ‘A kind of biotechnology involving manipulation of DNA is: ) © Ingenctic engineering, a @ ©) © @ DNA replication Denaturation (b) Genetic engineering (@) Renaturation chimerais ‘an enzyme that links DNA molecules. a plasmid that contains foreign DNA. ‘a virus that infeets bacteria, a fungus,— {iss 19. Bacteria protect themselves from viruses by fragmenting Viral DNA upon entry with (@ Methylase (b) Endonucleases (©) Ligases (d) Exonucleases 20. Restition enzymes are usedin genetic engineering because (@)_theycan join diferent DNA fragments (©) they can cleave DNA ata specific target. (©) they are nucleases that cut DNA at variable sites, (©) theyare proteolic enzymes which can degrade harmful enzymes, 21. Which ofthe followings is known as specific molecular scissors? @) Ligase (b) Helicase (©) Restriction endonuclease (@ DNA polymerase 22, The first restetion endonuclease reported was: (@) Hind (&) EcoRI (@) Hind (4) Bamttt 23. Restriction enzymes belong to a larger class of enzymes, which scalledas: (@ Ligases (b) Kinases (©) Nacleases (4) Polymerases 24, Restriction endonuclease - Hind II always cuts DNA molecules ata particular point by recognising a specific canal Critical Thinktng @) Sixtase pairs (b) Five bse pairs (©) Four bse pars (4) Seven basepairs 25. The sticky ends of DNA are due to (@) Unpaid bases (b) Endonucleases (© Freemethylaion _(@) None of these 26. Electrophoresis is used to (@) separate fragments of DNA, (©) clone gens. (©. eM DNA into fragments. (@_matcha gene with its function 27. Vectorsinclode (@) Bacterial plasmids (b) Viruses (©) Artificial chromosomes (€) All of these 28. The RTF region enables the plasmid to Critical Thinking (@)_ be wansmited to other bacteria by conjugation (b) undergo transformation. (© replicate inthe host el (None ofthese 29. _Arestiton fragment containing a specific ene of intrest can be identified by gel electrophoresis followed by transfering the DNA toa membrane as slid supprt matrix using a procedure called (@ Gencampiiication (©) Southern bot (©) Polymerase chain reaction (@) Western blot Cee 30, _Imexder for a prokaryote vector tobe propagated in a host bacterial cl the vector needs (@ Telomere (b) Centromere (©. Drug resistant genes (@ Anoriginofreplication 31. Revombinant DNA molecules were fis ofall sythessed by Paul Berg using DNA of (@) TMV and Salmonella (b) VirusSV-40and col (©) Influenza virus and Diplococcus bacterium (TMV and Agrobacterium tumefaciens 32, Which ofthe following is ota genetie vector? (a) Plasmid (b) Phage (c) Cosmid (d) Virusoid 33, A shuttle vectors @ puc (& pprs22 © Ye @ Allofthese 34. Plasmids are suitable vectors for gene cloning because (a) these are small circular DNA molecules which can integrate wih host chromosomal DNA. (©) thesearesmall ical DNA molecules with their own replication origin site (©) these can shuttle between prokaryotic and eukaryotic cells. (@ these often carry antibiotic resistance genes 435. Who created the first recombinant DNA molecule? (a) Nathan, Arber and Smith (b)_ Watson, Crick and Wilkins (©) Boyer and Cohen (@)_ None of the above 36. __vectors are used to clone DNA fragments more than 2500Kb, @ BAC) YAC) © MB @ puc 37. Restriction endonuclease cleaves the DNA molecule by hydrolysing: (@) H-bonds (b) Phosphodiester bonds (©) OH bonds (@) Phosphate bonds 38. Cosmids can clone DNA fragments upto: (a) ASkbin size (0) SOkbin size (©) 85kbinsize (@) 75kbin size ‘The tumour inducing capacity of Agrobacterium tumefaciens is located in large extrachromesomal plasmid called: (a) Riplasmid (0) Lambda phage (©) pBR322 @ Ti plasmid 40. Vent polymerase enzyme used in PCR is isolated from: (2) Thermococcus litoralis (b) Thermus aguaticus (© E.coli (@) Salmonella typhimurium 39.Biotechnology: Principles and Processes 41, Complementary base pairing is important for (@) ligation reactions with blunt end DNA molecules. (©) hybridisation between DNA and transeription factors. (©) restriction endonucleases to cut cell walls. (@)_ synthesising eDNA molecules from mRNA templates, 42, Cloning means that (@) all of the cells are derived from one cell and are genetically identical (©) a gene from one organism has been inserted into « ‘vector and successfully introduced into a host cell. (©) allofthe cells in a particular organism are identical (@)_Allofthe above 43, Recombinant DNA ean be transferred into host cell by (@) growing the host cell in growth medium containing ampicillin, coating the DNA with carbohydrates so thatthe cells, will engulf the DNA. (©) treating cells with calcium ions or electrical pulses to increase cell permeability (@)_ injecting proteins into host cells to make them more permeable, Expression vectors are different from other vectors because they (@) contain drug resistance markers. (©) contain telomeres. (©) contain regulatory regions that permit the cloned DNA to produce a gene product. (@)_ contain DNA origins. 45, Agenomic libraryis (@) where you look to find out how to make recombinant DNA. a listing of the known nucleotide sequences for a particular species. (©) all the genes contained in one kind of eel. (@)_acollection of cloned DNA pieces from an organism's ) 44. ) genome, 46. Which of the following enzyme is used in case of fungus to release DNA along with other macromolecules from the cells? (@). Lysozyme (b) Calulase (©) Chitinase (@) Amylase 447, Imagine gl through which DNA fragments have moved in response to an applied electrical current The band on tis gel thats farthest from the top (thats, from the place where the DNA fragments were added to the “well") represents the (a) shortest fragments of DNA. i (b) longest fragments of DNA. Uc Thinitng (©) restrition enzyme uscd to cut the DNA int fragments (@)_Tigaseused to bind the DNA fiagments together. 48, A biologist intends to use a polymerase chain reaction to perform a genetic task. The biologist probably is trying to (a) discover new genes. (b) clone a gene. (©) cutDNA into many small fragments (@) isolate DNA from a living cell 49. 50. si. 82, 55. 5. In genetic engineering, genes can be inserted from one organism into another or back into the original organism. using which of the following techniques? (a) Polymerase chain reaction () Gene gun (©) DNA hybridisation (@)_ Gel electrophoresis Which of the following is not necessary to execute @ polymerase chain reaction successfully? (@) All four DNA bases (b). Short DNA base primers (©) DNApolymerase (a) DNAlibrary It is now possible to breed plants and animals of desired Sec Critical Thinking (a) Tissue euture (b). Geneticenginecring (0 Mutational breeding (8) Chromosomeenginecring ‘Themain technique involved in agricultural biotechnology iscalled: (@) Tissue culture (© Plancbreeding Megaplasmids are found in (@ Ecol (6) Agrobacterium tumefaciens (© Pseudomonas (@ Bacitus substi Problems in obtaining large amounts of proteins encoded by recombinant genes ean often be overcome by using (@ BACs (b) Expression vectors (© YAcs (@) All ofthese “The technique in which foreign DNA is precipitated over surface of metal particles for passing into target ellis (@)_ Microinjection (b) Electroporation (© Particle gun (Chemical mediated gene transfer Gene gun ean introduce genes into cells withthe help of (@) Plasmids (b) Cosmids (©) Microscopie pels _(@) Phagemids Cloning oa gene involves (@) restriction endonucleases and ligase (b) plasmids and bacteriophage lamba (©). yeast artificial chromosomes and complementary base pairing (@ Allofthe above Polyethylene glycol method is used for (@) biodiesel production. (&) seedless fut production (@) energy production from sewage (@) gene transfer without vector. (b) Transformation (@ DNArepication Critical Thinkingoa0) (1040) 59. DNA cloning is beneficial because a cloned gene can be (@) sequenced using automated sequencing machines (@) transferred to other cells ofthe same organism, (©) transfered to cells in other organisins. (@ Allofthe above ‘The introduction of T-DNA into plants involves (@) exposing the plants to cold fora bref period (6) allowing the plant rootsto stand in water. (©) infection ofthe plant by Agrobacterium wmefaciens. (@) altering the pH ofthe soil, then heat-shocking the plants. AA dicotyledonous plant forms crown gall when (@) Agrobacterium tumefaciens comes in contact with the plant. Agrobacterium rhizogenes comes in contact with the plant. (© 2 specific part of DNA from the Ti plasmid gets integrated within the plant chromosome. 4 specific part of DNA from the Ri plasmid gets integrated within the plant chromosome 62. Due to chloramphenicol resistance gene, one is able to select transformed cell in the presence ofchloramphenicol ‘The chloramphenicol resistance gene in this case is called (@) Origin ofreplication (6) Cloning sites (© Selectablemarker_(@) None of these 63. Significance of heat shock method in bacterial transformation facilitates (@) binding of DNA tothe cel wall with thehelp of proteins. (©) uptake of DNA through membrane transport proteins. (©) uplake of DNA through transient pores in the bacterial cell wall (8) expression of antibiotic resistance gene. (4, Inmicroinjection method, the DNA is incorporated into host cell by using: (@) Micropipettes (©) Microncedles (©) High voltage electric impulse (@ Both a) and (b) 65. Bacterial cells are made competent by treating it with (@) divalent cation of calcium, () monovalent cation of calcium. (©) divalent cation of potassium. (@) monovalent cation of potassium. Purified DNA is precipitated by: (@) Chilled ethanol ©) Chilledacetone (©) Chilled methanol) Chilled butanol 667. Bleetroporation in cloning is (@) introduction of genetic material into transgenic animal (©) introduction of genetic material through viruses. (©) manipulation of cells by exposing them to a strong electric field (©) injecting DNA into the egg. (68. Thermal cycler is used in which one of hese reactions? @) Radioactivity (©). Enzyme catalysed reactions 60. ) @ 66. 69, 0. n, n B. 4, . 16. 5) (©) Chemical reactions (@ Polymerase chain reaction ‘Temperature required for various steps of polymerase chain reaction (A, B and) are: Critical Thinifng A. Denaturation B Annealing C Extension @ A-98°C, B-60°C, (6) A-40°C, B-72°C, © A-2°C, B-98°C, C-40°C (@ A-90°C, B-72°C, C-40°C AT, bacteriophage has a gene for the enzyme, lysozyme. ‘The function of this enzyme is to digest: (@ Cellwall (b) Cellmembrane (©) Golgi apparatus @ Plasmid Plants in comparison to animals are more rapidly ‘manipulated by genetic engineering. Select out the most probable reason for this. (8) Totipotency shown by plant cells () Presence of ell wall (©). Genetic engineering supplemented with plant tissue culture techniques (@ Both (@) and () Most sensitive technique to detect malignant cell in non- Hodgkin’s lymphoma is: (@)_ Polymerase chain reaction (©) Gene therapy (©) Stem cell therapy (@ None of the above Which one of the following pairs of terms/names means ‘one and the same thing? @) Gene pool of an organism — genome (6) Codon — gene (©) Cistron triplet (@_DNA fingerprinting ~ DNA profiling A bioreactor is: (@)_ fermentation tank. ©) culture containing radioactive isotopes. (© culture for synthesis ofnew chemicals. (@ hybridoma technology. Which of the following statements is not true for stirred tank fermentaion ? (@) Buffer needed to control pH (6) Batch and feed possible (©) Control dissolved oxygen (@ Easy in process sampling [At which stage after fermentation, its desired product is screened and purified” (@ Scaling (©) Screening (© Downstream processing (@ Batch culture c-2°C c-94°C— (Biotechnology: Principles and Processes {ost} —— @ EXERCISE-2| Picture Based MCQs 1. The image below represents the process of recombinant DNA technology with some tools and processes marked as A,B, Cand D. Which one of the following options is correct Iabeling for A,B, C and? Vectr, Foreign DNA DNA “OD Sanoons (plasmid) 4A +8 — jointrign CY DNA to plasmid 3. (@)_A—Exonuctease; B -Endonuclease; C—DNA ligase; D~ Transformation (©) A~Endonuclease; B~ Exonuclease; C~DNA ligase; D~Transformation (©) A—Exonuclease; B ~ Endonuclease; C — Hydrolase; D~ Transduction (@ A~ Restriction endonuclease; B ~ Restriction endonuclease; C - DNA ligase; D Transformation 2. ‘Theimage below represents polymerase chain reaction with, some steps marked as A, B, C and D. Identify the correct 4. labelling for A,B, C and D. (@)_ A~ Denaturation, B — Annealing, C ~ Extension, D—Amplified DNA. (b) A~ Annealing, B ~ Denaturation, C — Extension, D-Amplified DNA. (©) A~Denaturation, B~Annealing, C—Amplified DNA, D~Extension (@ A~Annealing, B—Denaturation, C-Amplified DNA, D-Extension ‘The image below represents a step in the formation of recombinant DNA (rDNA) between DNAS marked as A and B, Identify the correct labelling for A and B and also the enzymes involved in the formation of rDNA. ADNA BDNA PRG \ F ~TADNA [BDNA | Enzyme [Enzyme joining recognising |the sticky ends palindrome (@| Vector | Foreign | DNA ligase |EcoRT |()| Vector | Foreign | EcoRI — |DNALigase \@)| Vector | Foreign | Exonuclease|DNA ligase \(d)| Vector_| Foreign | DNA ligase |Exonuclease |dlentify the correct match for the given figure. Wells Smallest Largest DNA polymerase (Tag polymerase) + deoxymucleotides *| _ SER” 5 Wee 2 EEEEEEEEET ESEEEEEEGEET “Technique Description (@)| Electrophoresis. | Differential migration of DNA | fragments | (&)) Column Separation of chlorophyll chromatography | pigments (©)| Genecloning | Technique of obtaining identical copies ofa particular DNA segment ora gene (€)| Microinjection | Technique of introducing foreign genes into a host cell1042} Biology ud’) EXERCISE-3 Statement & Matching Based Topic-wise MCQs 1, Match column-I with column-I and choose the correct option. Colaman Cokuman-I K EcoRt T Bacillus amyloliquefaciens B Bam IL Haemophilus influenzae Hindi UL Escherichia coli D._pBR322 @ A-MkB-1;C-1:D-1V ()) A-M;B-1,C-1V, D-II (© A-IV;B-1;C-I D-Il (@ A-1B-IV;C-IL, D-Ill 2. Match column-I with column-II and identify the correct option Cohan in jamin A> Restriction enzyme _Jumping genes B__Transposons TL. Cloning vehicle Bacteriophage UL Hind Mt D._Palindrome IN. MALAYALAM @ A-TLB-IC-1:D-1V (©) A-I;B-;C-1V;D-L (© A-INV;B-kC-ID-Il @ A-EB-IV;C-I; D-II 3. Match column-I with column-II and select the correct combination from the option given below. Colamn-T ‘okanan-TT AL Host cells TL Thermus aquaticus B_ TagDNApolymerase IL An antibiotic CC Ampicitin Il, Microinjection D._Ethidium bromide __IV_DNAstaining (@ A-ILB-,C-1:D-1V (b) A-IB-I:C-1V;D=IL (© A-IN;B-L,C-I; D-IL @ A-EB-1V;C-I;D-m 4. Which one of the following pairs isnot correctly matched? (@ Plasmid — Small piece of extrachromosomal DNA in bacteria (©) Interferon — An enzyme that interferes with DNA. replication (©) Cosmid~ A vector for carrying large DNA fragments into host cells, (@_ Myeloma ~ Antibody-producing tumour cells, 6 10. Which ofthe following statements about restriction enzymes isincorrect? (@) They work on DNA extracted from all types of organisms. (0) Theyare used to glue together short segments of DNA, (©) They come in many varieties, each with its own DNA target sequence, (@)_Theyarehighly specific for their DNA target sequences. Match column-1 with column-II and select the correct answer using the codes given below, T Cohunnn-T A. Tiplasmid T Agrobacterium ‘tumefaciens B Salt Tl Cancerous cells C_ Retrovirus Il, Recombinant DNA D._Ligase IV. Restriction enzyme @ A-W;B-1,C-1D-I () ACILB-1C-1V;D-Ill (© A-IV;B-1;C-I; D-IL (@ A-LB-IV;C-I;D=IIL Which of the followings incorrectly matched? (@) Agarose — Sea weeds (©) Thermus aguaticus — “TEDNA’ (©) Plasmid DNA Vector @ Salt Restriction endonuclease ‘Which of te following is/are part(s) of biotechnology? (Invitro fertilisation (i) Synthesis of a gene (ii), Correcting a defective gene (iv) Developinga DNA vaccine @ @adGi ©) Gidanddi) (© Gi,Giiyand Gv) (© GG.) andy ‘Which of the following is/are used in recombinant DNA technology? (Agarose gel (i) Ethidium bromide (ii) Plasmid veetor (iv) Restriction endonuclease (@) @and Gi) (©) Gand it) (©) Giiyand iv) @ @,Gi),Cii) and iv) Which of the following statements are correct about scientist Paul Berg? ()Heis the Father of genetic engineering. (i) Hetransferred gene of SV40 virus into E. col. (ii) He got the Nobel Prize in chemistryin 1980, (iv) He developed hybridoma technology for the production of monoclonal antibodies. @ @andGi) (©) (and (ily (© Giiyand div) © (,Gi),and Giiy——{ Biotechnology: Principles and Processes | 2. 13. Molecular probes used for identification of recombinant |. clone carrying the desired DNA insert can be (@ denatured double-stranded DNA probes. (double-stranded RNA probes. (i) protein probes. (iv) single-stranded DNA probes. (@) @and (ii) (©) Giana) © @and (iv (@ (,Gi, (ii) and Gv) ‘Match the column-I and column-lII and select the correct option. Column-T Cohan Cos ‘Gene that can be constructed in the laboratory, ifits base sequence can be deduced from the amino acid sequence of the A) Radioactive gene B. Artificial gene 1 to identify colonies of genetically engineered bacteria that makea particular gene product. ‘Abnormal enhanced replication of@ plasmid results in many copies 16 ofthe plasmid in each cal Allarge population of identical cells. The use of entire array of genes of an organism in order to obtain the particular gene product. @ A-IB-1,C-ii, D-IVE-V () A-LB-ICAD-IV;E-V (©) A-LB-I;CAV;D-IGE-V Amplification IL D. Toproduce clone IN. E Shotgun cloning = W. (@) A-I;B-Ill,C-V;D-1V;E-1 ‘Match the following columns and select the correct answer using the codes. ‘Column ‘Colunnn-MT 1, K_ RecombinantDNA I Sea weeds B Ethidium bromide 1. DNAstaining C_ Gelelectrophoresis MIL Plasmid DNA that has incorporated human DNA D. Agarose IN. Process by which DNA fragments are separated based on their size @ A-M;B—1,C-1,D-1 (o) A-II;B-Il,C-I;D-IV © A-I;B-1,C-IV;D-I @ A-M;B-1V;C-1;D-11 1043) Identify the incorrect statement. (@) Mobile genetic elements, transposons were visualised byBarbara McClintock Udder cell and somatic cell are used to produce the cloned sheep by nuclear transplanation method. (© In pedigree analysis, a person immediately affected by or concerned with an action is called propositus. (@)_ DNA ligases are used to cleave a DNA molecule. Process of Recombinant DNA Technology © ‘Match colum-I with column-II and select the correct answer using the codes given below. ‘Cohan Coaman-TE Recombinant T Vector DNA technology B Cloning IL Sealingenzyme vehicles C. Macromolecular MIL Electrophoresis, separation D._DNAligase IV. Genetic engineering @ A-W;B-[.C-1D-mr (©) A-I;B-1;C-1V;D-mL (© A-IV;B-1,C-I;D-I (@ A-bB-1V,C—I D-IL Match column-{ with column-II and select the correct answer using the codes given below. ‘Column Coame-T Primers T PCR B Separation and 1. C,H,0H purification of| products Precipitation of I, Uptake of foreign DNA. DNA by bacterium D._Transformation IV_Dowmstream processing @ A-WB-EC-hD-m (©) A-IbB-C-1V;D-IIT (© A-IV;B-1,C-IlkD-Il (@ A-LB-1V;C-I;D-Iml Match column-I with cofumnn-II and select the correct answer using the codes given below. Cohannn-l ‘Cohamnn-T x PCR T Large scale culture B Bioreactor Tl, Toinducealien DNA in host cell © Genegun I, Restriction endonuclease D._EcoRL IN._Amplification of gene @ A-IV;B-1;C-I D-Ill () A-I;B-1;C-IV;D-I01 (© ACIV;B-1;C-I D-Il @ A-EB-IV;C-I D-Ill(foaa. { Biology )- 18. Match column-I with column-I and select the correct answer i) Cohen and Boyer are known as Father of genetic using the codes given below. engineering Cohan Coane Gi) When eutby the same restriction enzyme, the resultant Phsmid T_ Selecablemarker DNA fragments have the same kind of sticky ends and B. amp 1 Extrachromosomal DNA these ean be joined together using DNA ligase © Tiplasmid TM. Enzyme + iv) Endonucleases remove nucleotides from the ends of D. Chitinase WV. Agrobacterium the DNA whereas exonuleases make cuts at specific tumefaciens positions within the DNA. @ A-WB-kC—1D—I (0) Presence of more than one recognition sites within the (©) A-IB=1;C-1V;D-IIL vector will generate several fragments which will (© A-IB-RC=IkD =I complicate the gene cloning @ A-iB-1V;,C-1;,D—m (i), Humulin was the frst recombinant DNA based product, 19. Match column with colimn-II and cect the corect answer produced and marketed in India using the codes given below. (i) YAC vectors contain the telomeric sequence, the ~~ Column-T (Column-Il centromere and autonomously replicating sequence K Agarose TPR fom yeast chromosomes. B Opines I. Genegun (il) Alkaline phosphatase is used to prevent unwanted self C_Biotisic ML, Tiplasmid ligation ofthe vector DNA molecules in procedures of D._Thermalyeter NV. Sea weeds DNA technology. (@ A-IV;B-M,C-1,D— x) pBR322 vector was the first artificial ideal vector (b) A-I;B-1;C-1V;D-m constructed by Bolivar and Rodriguez. (© A-W;B-EC-IkD-IL (8). Plasmid DNA is coated with histone proteins and can (@ A-kB-1V,C-1;D-m act as genetic factor. 20, Match column-Iwithcolum-I andselectthecorrectanswer (ip, (vi)and(x) winged gen es © Gama A. Southern blotting 1 Running of DNA 2 OED SS Cin) ent) fragments on gel (4) i), Gi), (iv), (vi) and (x) D Gieeemen! 1 icaaraain 22. Match the following columns and select the correct answer ashi using the codes given below. C Cleavage TL Transfer of DNA ze Comme ipl dn gal so 1K Bacterial cellisteated Lysozyme nitrocellulose disc with D._ DNApprobing IV. Searching for desired B._Plantcellistreated 1M, Cellulase DNA fragments with (@) A-IV;B-LC-ID-It Fungal cell is treated IHL Chitinase (&) AI; B-I;C-IV; D-IL with, (© A-IB-RC-kD-I a (@) A-1,;B-IV;C-I, D-II 21, Tdenify the correct statements @) m} om) 1 (@ The first recombinant DNA was constructed by using. 1 piece of DNA from a plasmid carrying antibiotic resistance gene in the bacterium Salmonella ‘typhimurium and linked it tothe plasmid of E. col. (| uo} m | 1 fo) 1} a | m | mj 1 |(Biotechnology: Principles and Processes — ~ @ EXERCISE-4| Assertion Reason MCQs| Directions : These questions consist of two statements, each printed as Assertion and Reason. While answering these ‘questions, you are required to choose any one of the following four responses. (a) Ifboth Assertion and Reason are correct and the Reason is, correct explanation ofthe Assertion. Ifboth Assertion and Reason are correct but Reason is not 4 correct explanation ofthe Assertion. (©) Ifthe Assertion is correct but Reason is incorrect. (@ Ifboth the Assertion and Reason are incorrect. (O) 1. Assertion sequence. Reason: Palindromic sequences read the same in both directions of the two strands. 2. Assertion: Restriction enzymes Hind I and Hpa are produced from two different genera of bacteria, + Restriction enzymes recognise palindromic Reason: Ligases produce thenick in the recombinant DNA molecule. ‘Assertion : The uptake of DNA during transformation isan active, energy requiring process. Reason : Transformation occurs in only those bacteria, which possess the enzymatic machinery involved in the active uptake and recombination. ‘Assertion : In combinant DNA technology, human genes are often transferred into bacteria (prokaryotes) or yeast (cokaryore), ‘Reason : Both bacteria and yeast multiply very fst to form ‘huge population which express the desired gene. ‘Assertion: Electroporation uses electric pulses for making competent host. Reason: The electrical pulses induce transient pores inthe plasmalemma through which DNA molecules are Reason: Hind isproduced ftom Haemophilis while Hpa __‘neoPorsted. is produced from Hematococeus, or eaemomaes ptenstrnkc saa ‘bright orange coloured bands. 3. Assertion : Restriction enzymes of different organisms that recognise the identical sequences are called isoschizomers. Reason : They are present only in eukaryotes, 9. Assertion: Genetic engineering requires both nucleases and ligases. Reason: The separated DNA fragments can be seen after staining the DNA with compound EtBr. 4. Assertion: Agrobacterium faciens is a 4: Absertoa: Rest isiondigrtion wa proces awtingDNA ‘41 Asertions Agrobaciarien|feraefactens is spashowen of ijieariaios eases. several monocot plants. Reasoa: DNA tgaseoins two DNAS. Reason: Retroviruses in plants have ability to transform 5. Assertion: Restriction endonucleases are also called ___‘Darmal cells into cancerous cell ‘molecular scissors 15. Assertion: Downstream processing is generally considered Reason: When fragments generated by restriction __-‘oredificultand costlier in plants than thatin micrebos, ndanueleaen are mined, they join together du to their R800: Rhizoscrtion is use as a method to facilitate sticky ends easier recovery of recombinant protein from plants. 6. Assertion: Abacteral cell with no restriction enzymes will Assertion : Superbug has been biopatented on the name be easily infected and lysed by bacteriophages. of Prof. AChakrabarty. Reason: Restriction enzymes catalyse synthesis of Reason : Superbug was created by him with exceptional protective coat around bacterial cell that prevents degradative plasmid to degrade toxic substances. bacteriophage attack. 17. Assertion : One application of genetic engineering is the 7. Assertion: In gel electrophoresis, DNA fragments are __ Production of human insulin by microbes. separated Reason : Gene for production of human insulin ean be Reason: DNA is negatively charged, soit moves towards _‘transferred to Escherichia coli by recombinant DNA anode under electric field. technique. Assertion: All endonuclease cut DNA at specific sites. _‘'8- _Assertion : All GM organisms arerich in chimeric DNA Reason : Endonueleases are found in viruses (DNA) ‘Reason Chimeric DNA constitutes genes of two different organisms.a Past Year MCQs (aipmT/NeeET & alims)| (eo TENERGISE-5] AIPMT/NEET DNA precipitation out of mixture of biomolecules can be achieved by treatment with: 12019) (@) Isopropanol (b) Chilled ethanol (©) Methanol at room temperature (@) Chilled chloroform, Which one of the following equipment is essentially required for growing microbes on a large scale, for industrial production of enzymes? 2019) (@) BOD incubator (b) Sludge digester (©) Industrial oven (@) Bioreactor Following statements describe the characteristics of the enzyme Restriction Endonuclease. Identify the incorrect statement. [2019] (@)Theenzyme cuts DNA molecule at identified position \within the DNA. ‘The enzyme binds DNA at specific sites and cuts only cone of the two strands. ‘The enzyme cuts the sugar-phosphate backbone at specific sites on each strand. The enzyme recognises a specific palindromic nucleotide sequence in the DNA. The correct order of steps in Polymerase Chain Reaction (PCRs: (2018) (@) Extension, Denaturation, Annealing (b) Annealing, Extension, Denaturation (©) Denaturation, Annealing, Extension (@)_Denaturation, Extension, Annealing ‘Which of the following is commonly used as a vector for introducing a DNA fragment in human lymphocytes? ©) © @ 2018] (@) Retrovirus (©) Tiplasmia (©) ppR322 @ A phage ‘The process of separation and purification of expressed protein before marketing is called: 2017], (@ Downstream processing (6) Bioprocessing (©) Postproduction processing (@) Upstream processing What is the criterion for DNA fragments movement on agarose gel during gel electrophoresis? 017) (@) The smaller the fragment size, the farther it moves. (©) Positively charged fragments move to farther end, (©) Negatively charged fragments do not move. (@)_Thelarger the fragment size, the farther it moves. A gene whose expression helps to identify transformed cll is known as: (2017) (@) Vector (© Structural gene (©) Plasmid (@) Selectable marker % 10. 2. 1B. 14, 16. "1. 18. 19, ‘The DNA fragments separated on an agarose gel can be visualised after staining with: (2017 (@) Acetocarmine (©) AnitinebIue (©) Ethidium bromide _(@) Bromophenol blue ‘Which of the following is #resirieion endonuclease? 2016) (@) Hindi (b) Protease (©) DNase @ RNase ‘The Tag polymerase enzyme is obtained from: [2016] (@) Thermus aquaticus (b) Thiobacillus ferroxidans (©) Bacillus subiilis (@ Pseudomonas putida ‘Which of the following is not a feature ofthe plasmids? (2016) (@) Independent replication (©) Circularstructure (© Transferable @ Single-stranded ‘The DNA molecule to which the gene ofinterestis integrated for cloning is called: [2015] @ Vector (6) Template (©) Carer (@ Transformer ‘The cutting of DNA at specific locations became possible with the discovery of: [2015] (@) Probes (b) Selectable markers (© Ligases (@) Restriction enzymes ‘Commonly used veetors for human genome sequencing are: (2014) (@) TDNA (b) BACand YAC (©)_Expression vectors. (@) T/A cloning vectors. ‘Which vector can clone only small fragment of DNA? (a) Bacterial antificial chromosome 2014) (b) Yeast artificial chromosome (© Plasmid @ Cosmid In vitro clonal propagation in plants is characterised by: (@) PCRandRAPD 2014) (©) Norther blotting (©) Electrophoresis and HPLC @ Microscopy An analysis of chromosomal DNA using the Southern hybridisation technique does not use: 2014) (@) Electrophoresis (©) Blotting (©) Autoradiography (@) PCR Genes of interest can be selected from a genomic library by using: (2013) (@) Restriction enzymes (6) Cloning vectors (© DNA probes (@) Gene targetsHind 20. 21. 2, 23. 24, During the process of isolation of DNA, chilled ethanol is added to: (2013) (@) Remove proteins such as histones (b).Precipitate DNA (©) Break open the cello release DNA (@ Facilitate action of restriction enzymes ‘The colonies of recombinant bacteria appear white in contrast to blue colonies of non-recombinant bacteria because of, [2013] (@) Insertional inactivation of alpha-galactosidase in non- recombinant bacteria, (b) Insertional inactivation of alpha-galactosidase in recombinant bacteria (©) Inactivation of glyeosidase enzyme in recombinant bacteria (® Non-recombinant bacteria containing beta- galactosidase DNA fragments generated by the restriction endonucleases in a chemical reaction can be separated by: 2013] (@) Polymerase chain reaction (©) Electrophoresis, (©) Restrition mapping (©) Centrifugation ‘The figure below shows three steps (A, B, C)of Polymerase ‘Chain Reaction (PCR). Select the option giving correct identification together with what it represents. [2012S] Region to be amplified (@) B~ Denaturation at temperature of about 98°C separating the two DNA strands (©) A~Denaturation ata temperature of about SO°C (© C~Extension in the presence of heat stable DNA polymerase (@ _A~Annealing with two sets of primers ‘The figure below isthe diagrammatic representation ofthe E.coli vector pBR322. Which one of the given options correctly identifies its certain component(s)? [2012 M] 25. 26. a 28. 29, 30. 31 Bam ‘Sal Phu ori orginal restriction enzyme rop ~ reduced osmoti pressure (© Hind il, EcoRI—selectable markers (@_ amp®, tet® — anti resistance genes Which one of the following represents a palindromic sequence in DNA? [2012 S] @ S-GAATIC-3 3-CTTAAG-5' 5/-CCAATG-3" 3-GAATCC-5' 5/-CATTAG-3 3/-GATAAC- 3/-GATACC-3 3)-OCTAAG- In genetic engineering, the anibiotis are sed (@) asselectable markers. (©) to select healthy vectors (©) to keep the cultures fre of infection. (@) as sequences from where replication tarts Biotistes (gene gun is suitable for (@) DNA fingerprinting (b) disarming pathogen vectors (©. transformation of plant cells (@)_ constructing recombinant DNA molecules. For transformation, microparticles coated with DNA to be bombarded with gene gun aremadeupof: (2012 (@ SilverorPlatinum (6) PltinumorZine (© Siiconor Platinum —_(d) Gold or Tungsten ‘Which one isa true statement regarding DNA polymerase used in PCR?’ 2012] (@)_Itisusedtoligate introduced DNA in recipient cel (b) serves as selectable marker. (9. Itisisolated from a vius (@)_Itremains ative at high temperature. (A single strand of nuclei acid tagged with a radioactive @ © © © @ (20128) (2012 My) ‘molecule is called: [2012] (a) Vector (b) Selectablemarker (©) Plasmid @ Probe PCR and Restriction Fragment Length Polymorphism are the methods for: [2012M] (@) Study ofenzymes _(b) Genetic transformation (© DNAsequencing __(@) Genetic fingerprinting36. 38. 048 ‘Agarose extracted from sea weeds finds use in: [2011] (@) Spectrophotometry () Tissue culture (© PCR (@ Gotelectrophoresis Which one ofthe following techniques made it posible to genetically engineer living organism? oum} {@) Recombinant DNA techniques (©). X-ray diffraction (©) Heavier isotope labelling (@) Hybridisation Theresa restrition endonuclease called EcoRI. What does co part init stand for? (2011) @ Colon (b) Coctom (€) Coenzyme (a) coli Stirred-tank bioreactors have been designed for [2010] {@) addition of preservatives to the product (©) purification ofthe product. (©) ensuring anaerobic conditions inthe culture vessel (@)_ availablity of oxygen throughout the process. DNA or RNA segment tagged with aradioactivemolecule is called: 2010] (@) Vector (b) Probe (©) Clone (4) Plasmid Which one ofthe following is used as vector for cloning ‘genes into higher organisms? 2010) (a) Baculovirus (©) Salmonella typhimurium (©) Rhizopus nigricans (@) Retrovirus Restriction endonucleases are enzymes which [2010] (@) make cuts at specific positions within the DNA molecule, (b) recognise a specific nucleotide sequence for binding ofDNA ligase. (©) restrict the action of the enzyme DNA polymerase. (@)_removenucleotides fiom the ends ofthe DNA molecule. Which one of the following palindromic base sequences in DNA can be easily cut at about the middle by some particular restriction enzyme? Peo10) @ OGTTCG vom ATGGTA... © .GATATG. CTACTA... © .GAATTC. CTTAAG... @ sm CACGTA... .CTCAGT. SIMPLE MCQs In gene cloning: [2019] (@) Gene is isolated and inserted in the same organism, (b) Gene is isolated and inserted in different organism. (©) Gene is isolated and inserted in plasmid of other organism. (@) Gene is isolated and inserted in chromosomal DNA. Which of the following statement is not correct about cloning vector? * 12017] —_ = (©) ‘Or iasoquence responsible fr etoling te ony umber ofthe inked DNA (2) Seetablemarkers selectively pemitthe growth ofthe nonransformants, (©) In orde oink te alien DNA, the vector needs to have single recognition ste fo the commonly used restristion enzyme, (€)Theligation often DNA scartiedout sta estition stepresentin onc ofthe wo antibnoe eisance genes, 42. Which oneo the filloving palindromic tase sequen in DNA can be easily cut at about the middle by some particular restriction enzyme? 2016) Senn OGTTCG in ATGGTA oon 5 AGATATG ond CTACTA... 5 GAATTC. 3 CTTAAG 5) LEACGTA con 3) CTEOAGT on! 49, Choose the correct option. 2015] Cla. Hind A B € D @ [Hind | BeoRt | amp® | ori (| Hind — | Bambtt | kan® | amp® (© [Bama | Pst ori amp @{zeoRt | BamHt | amp®| ori 44, Genes offnterest can be selected from a genomic library by using: [2014] (@) Restriction enzymes (b) Cloning vectors (© DNA probes (@ Gene targets 2 Sterile Air=F Funalor “| at, Amerton: Agractriom tmefeclns i popula in \@) | Gene gun venes direct gene genetic engineering because this bacterium is associated en ee ih woof coisa uc crea © MEHL eee | enone saa A lea es ead em Jo Stir an captains jaca eae ere ocr oa cane ae Sieh the bacerun e eoce \(d) | Respirometer Finding out rate of 48. Assertion: A piece of DNA inserted into an alien organism 1 respiration generally does not replicate if not inserted into a dic AbegrCRcpantormmagiAmasaiocacteed (cae fomone DNA empatomoece? i201 Reason: Chromosomes have specific sequences called “ori” region where DNA replication is initiated [26 May, 2018] 49, Assertion: Insertion of recombinant DNA within the coding @ 4 ) 32 @6 @s Directions for (Qs.47-50): Each ofthesequesions containsan sequence of galactosidase results in colourless colonies. Acero lousy en Realm tly nd atone: eeneofer res innacatin ene question on he ass of following options. Youhavetoselest _p,exactosidaseknown as insertional inatvation. [20 the one that best des the two statements. Ca Ms faa eats oo rere ected °° Atertion: Inremombinaat DNA technology, man genes fete tere ecto are often transferred into bacteria (prokaryotes) or yeast (b) both Assertion and Reason are correct, but Reason is not (eukaryote. the correct explanation of Assertion. Reason: Both bacteria and yeast multiply very fast to form (©) IAssertion is correct but Reason is incorrect. hhuge population, which expresses the desired gene. [2015] (@)_Ifboth the Assertion and Reason are incorrect. "ANSWER KEYS Exercise 1: Tople-wise MCQs O12 lol7 o[sl@[slolal@] elo {s7[@]s]@[ sl @ @ [10 [© [as | @ [26 [@ [34 [om] 2 | @ | 50] @ | s8 | @ | 66 | @ | 4 | olulol»|®27l@[slolslols] om] s[@] 7 |o|s|o @lnl@l2|@)2{@|s|olalols|@ | ole || alo @ [3 [@ [21 | © [29 | | 37 [@ [4s To] 3 [| or | © | 09 | @ @)4l@]2l@[s|@|ss[ ola lols [@]e]o| 2] a @)sl@olalolsl|wm|[»|@la[@l[sloleslolul|@ @lil@lal@l2_@lol@lslm]sslolelol2lo Bxercise 2: Picture Based MCQs 11/@[?2 [3 lols lo Exercise 3: sment & Matching Based Topic-wise MCQs T]@]4 /6|7 [vl @[sl@[ol@[vtol[zlo mlslolslol[ulols/@l{@l{wle@ @l[sl@lsl@[elolslolslwmlalo Bxereise 42 Assertion Reason MCQS sTol[u lo] sy o[is[ofizto wl@lel@!sl@lel@lslo sst Year MCQs (AIPMIT/NBET & AIMS) z2T@l[4 [wots lo ifm «T@o[ulte@ a | @ [2] @] a] @ 36] om | 41 | & | 6 LO 2[@l7lo[2l@ zlolalolzl@l[sl@][a2loi|7|@ s/o! s[@[ alo 23 | © | 28 | @ [33 | @ | 38 | @ | 3 | @ | 8 | +lol>lolul@ 2 | @] 29] @] 34 [@]s | o | 44 [ol | @ s[@lol@lslo 2 [@ |30|@] 35] @/ 4 | wm [4s Lo [slo