201. Why are low-copy-number plasmids generally less preferred when a cloned DNA fragment must be amplified rapidly inside a host cell?
ⓐ. They cannot carry selectable markers.
ⓑ. They cannot replicate in bacterial hosts.
ⓒ. They always contain many restriction sites.
ⓓ. Fewer plasmid copies produce fewer linked copies of the inserted DNA.
Correct Answer: Fewer plasmid copies produce fewer linked copies of the inserted DNA.
Explanation: The foreign DNA replicates along with the plasmid vector to which it is attached. When the plasmid exists in only a few copies per cell, the inserted DNA also remains in relatively low copy number. This reduces amplification efficiency when many copies are needed quickly. High-copy-number vectors are therefore often preferred for cloning purposes.
202. Which screening method identifies recombinants by disrupting a coding sequence so that colony color changes on a chromogenic medium?
ⓐ. Ori-based copy-number selection
ⓑ. β-galactosidase-based color screening
ⓒ. Agarose gel electrophoresis
ⓓ. Calcium chloride competence testing
Correct Answer: β-galactosidase-based color screening
Explanation: In this approach, insertion of foreign DNA disrupts the coding sequence for β-galactosidase. Colonies carrying non-recombinant plasmids remain blue on chromogenic medium, whereas recombinants appear colorless. This makes screening possible without relying only on antibiotic-resistance loss. The method is based on gene disruption causing a visible colony-color difference.
203. Which observation most strongly suggests that bacterial cells were made competent before transformation?
ⓐ. Their DNA bands became visible under UV light.
ⓑ. Their chromosome was cut at palindromic sites.
ⓒ. Their ability to take up external DNA increased markedly.
ⓓ. Their plasmids changed into linear RNA molecules.
Correct Answer: Their ability to take up external DNA increased markedly.
Explanation: Competence refers to the condition in which bacterial cells can take up DNA from their surroundings more effectively. It is produced experimentally because DNA does not normally enter bacterial cells easily. A rise in DNA uptake is therefore the direct sign of competence. The other options describe unrelated processes such as restriction digestion or gel visualization.
204. Which method is correctly matched with the problem it is mainly used to overcome?
ⓐ. Calcium treatment followed by heat shock — poor natural uptake of DNA by bacterial cells
ⓑ. Micro-injection — size-based separation of DNA fragments
ⓒ. Biolistics — purification of recombinant proteins
ⓓ. Elution — transfer of genes into animal-cell nuclei
Correct Answer: Calcium treatment followed by heat shock — poor natural uptake of DNA by bacterial cells
Explanation: Bacterial cells do not normally take up DNA efficiently because DNA is hydrophilic and the membrane acts as a barrier. Calcium treatment helps make the cells competent, and heat shock promotes uptake of the DNA. This combination is therefore used to assist transformation in bacteria. The other methods are used for different purposes.
205. After plant cell walls have been digested and cellular contents released, which step should logically occur before chilled ethanol is added during DNA isolation?
ⓐ. Direct transfer of DNA into a host cell
ⓑ. Removal of RNA and proteins by suitable enzymes
ⓒ. Restriction digestion of the purified DNA
ⓓ. Ligation of DNA with a plasmid vector
Correct Answer: Removal of RNA and proteins by suitable enzymes
Explanation: Chilled ethanol is used to precipitate DNA after major contaminants have been removed. Before that stage, RNA and proteins should be digested so that the DNA preparation becomes cleaner. Ribonuclease removes RNA, and protease removes proteins. Only then is ethanol precipitation used to recover relatively purified DNA.
206. A PCR mixture contains template DNA, primers, nucleotides, and buffer, but no thermostable DNA polymerase. What will happen during the extension stage?
ⓐ. No new DNA strands will be synthesized.
ⓑ. The primers will convert into restriction enzymes.
ⓒ. The template DNA will ligate to itself automatically.
ⓓ. Amplification will continue normally because heating is sufficient.
Correct Answer: No new DNA strands will be synthesized.
Explanation: DNA polymerase is the enzyme that actually extends the primers by adding nucleotides to form new DNA strands. Without it, the denaturation and annealing steps may still occur, but extension cannot proceed. As a result, the target sequence will not be amplified. Heating alone cannot replace enzymatic DNA synthesis.
207. Why are two primers used in PCR rather than only one?
ⓐ. One primer is needed for DNA, and the other is needed for RNA.
ⓑ. Two primers make DNA ligase unnecessary.
ⓒ. Two primers define the two boundaries of the DNA segment to be amplified.
ⓓ. Two primers are required to create sticky ends for cloning.
Correct Answer: Two primers define the two boundaries of the DNA segment to be amplified.
Explanation: PCR is designed to amplify a specific region rather than the whole DNA molecule. The two primers bind to opposite strands at the flanking regions of the target sequence, marking where amplification begins on each side. This lets repeated cycles copy the chosen segment efficiently. With only one primer, controlled amplification of the selected region would not occur in the normal way.
208. In a continuous culture system, fresh medium is added and spent medium is removed mainly so that cells
ⓐ. lose their plasmids more slowly
ⓑ. remain in active exponential growth for a longer time
ⓒ. stop dividing and store the final product
ⓓ. become resistant to contamination automatically
Correct Answer: remain in active exponential growth for a longer time
Explanation: Cells produce useful products most effectively when they are metabolically active and multiplying well. Continuous culture helps preserve that state by supplying nutrients and removing waste products. This delays the onset of unfavorable conditions that would slow growth. The main goal is to maintain a productive biomass for longer periods.
209. Which feature of a bioreactor allows culture material to be withdrawn during operation for checking contamination, growth, or product formation?
ⓐ. Agitator
ⓑ. Foam-control unit
ⓒ. Sparger
ⓓ. Sampling port
Correct Answer: Sampling port
Explanation: Monitoring is important during large-scale production because conditions can change over time. A sampling port allows small amounts of culture to be removed without disturbing the whole system significantly. The sample can then be used to assess contamination, cell density, or product level. This makes the sampling port a key monitoring feature.
210. A purified recombinant therapeutic product is unstable during storage. Which downstream step becomes immediately important?
ⓐ. Restriction digestion
ⓑ. Host transformation
ⓒ. Suitable formulation with stabilizers or preservatives
ⓓ. Primer annealing
Correct Answer: Suitable formulation with stabilizers or preservatives
Explanation: Even after purification, a biotechnology product may not yet be ready for practical use. If the product is unstable, formulation is needed to maintain its activity and shelf life during storage and handling. This may involve adding stabilizers or preservatives as appropriate. Formulation is therefore a post-purification step of downstream processing.
211. In the sequence “competent bacteria + recombinant plasmid + heat shock + plating on ampicillin medium,” which step directly distinguishes transformants from non-transformants?
ⓐ. Plating on ampicillin medium
ⓑ. Making the bacteria competent
ⓒ. Heat shock treatment
ⓓ. Mixing cells with plasmid DNA
Correct Answer: Plating on ampicillin medium
Explanation: Competence and heat shock help DNA enter bacterial cells, but they do not by themselves reveal which cells have taken up the plasmid successfully. Antibiotic plating provides that distinction because only cells carrying the appropriate resistance marker survive. This step acts as the selective filter. It therefore identifies transformants among the treated population.
212. Which pair is correctly matched with the function it directly performs?
ⓐ. Ethidium bromide — amplification of target DNA
ⓑ. DNA ligase — covalent sealing of joined DNA fragments
ⓒ. Chitinase — release of DNA from bacterial cell wall
ⓓ. ori — identification of recombinant colonies by color
Correct Answer: DNA ligase — covalent sealing of joined DNA fragments
Explanation: DNA ligase forms phosphodiester bonds between adjacent nucleotides and seals the backbone where two DNA fragments have been joined. This is essential after compatible ends have aligned during recombinant DNA formation. The other pairs are mismatched because ethidium bromide is used for staining, chitinase acts on fungal cell walls, and ori initiates replication rather than colony-color identification. Ligase is therefore the direct joining enzyme.
213. Which of the following would be classified under the broad meaning of biotechnology but not necessarily under its modern restricted meaning?
ⓐ. Preparation of bread using yeast fermentation
ⓑ. Introduction of a selected gene into a bacterial plasmid
ⓒ. Transfer of T-DNA into a plant cell using a modified vector
ⓓ. Amplification of a target DNA segment by PCR
Correct Answer: Preparation of bread using yeast fermentation
Explanation: The broad meaning of biotechnology includes the use of living organisms or their enzymes for useful products and processes. Bread making with yeast fits this definition because it depends on microbial activity. The modern restricted meaning is narrower and is centered on molecular manipulation such as gene transfer and recombinant DNA methods. Fermentation is therefore a classical broad example rather than a necessary example of restricted biotechnology.
214. Which statement best explains why bioprocess engineering is indispensable even after a useful recombinant organism has already been created?
ⓐ. It converts the cloned gene directly into a purified protein.
ⓑ. It replaces the need for vectors and host cells.
ⓒ. It provides sterile and controlled conditions for large-scale growth and product formation.
ⓓ. It cuts DNA at the same site in both vector and insert.
Correct Answer: It provides sterile and controlled conditions for large-scale growth and product formation.
Explanation: Creating a recombinant organism is only one part of biotechnology. To obtain the desired product in useful quantity, the cells or microbes must be grown under carefully controlled conditions at larger scale. Bioprocess engineering handles this requirement through sterility, nutrient control, and maintenance of appropriate growth conditions. It therefore supports production rather than genetic alteration itself.
215. In the first recombinant DNA experiment, which sequence of events is most appropriate?
ⓐ. E. coli was first chosen, then ligase was discovered, then plasmid DNA was prepared.
ⓑ. A plant vector was modified, then DNA was amplified by PCR, then transferred into yeast.
ⓒ. Restriction enzymes were used to stain DNA, then plasmids were purified, then protein was isolated.
ⓓ. An antibiotic-resistance gene was linked to a plasmid, and the recombinant DNA was then multiplied in E. coli.
Correct Answer: An antibiotic-resistance gene was linked to a plasmid, and the recombinant DNA was then multiplied in E. coli.
Explanation: The early recombinant DNA experiment depended on joining a selected gene to a plasmid that could act as a vector. After construction of the recombinant molecule, it was introduced into E. coli for multiplication. This sequence showed that DNA from different sources could be linked and maintained in a bacterial host. That was a crucial step in the rise of recombinant DNA technology.
216. In the name EcoRI, the letters "co" are derived from the
ⓐ. order of enzyme discovery
ⓑ. species name of the source organism
ⓒ. strain designation of the bacterium
ⓓ. nucleotide sequence recognized by the enzyme
Correct Answer: species name of the source organism
Explanation: Restriction enzyme names are built from the source bacterium in a systematic way. In EcoRI, "E" comes from the genus Escherichia, while "co" comes from the species coli. The following letter refers to the strain, and the Roman numeral indicates the order of isolation. The name therefore reflects biological origin rather than recognition sequence.
217. Which sequence pair is correctly judged as palindromic only when strand orientation is taken into account?
ⓐ. $5'$—GATATC—$3'$ and $3'$—CTATAG—$5'$
ⓑ. $5'$—GATTACA—$3'$ and $3'$—CTAATGT—$5'$
ⓒ. $5'$—AACGTA—$3'$ and $3'$—TTGCAT—$5'$
ⓓ. $5'$—GTCAAG—$3'$ and $3'$—CAGTTC—$5'$
Correct Answer: $5'$—GATATC—$3'$ and $3'$—CTATAG—$5'$
Explanation: A DNA palindrome is identified by reading both strands in the same orientation, not by simply checking whether they are complementary. In option A, the sequence reads the same when this orientation rule is applied. The other options may look related or complementary, but they do not satisfy the palindromic condition properly. The key point is orientation-based symmetry, not ordinary complementarity alone.
218. A DNA fragment is seen closer to the well than another fragment after agarose gel electrophoresis. The most reasonable inference is that the fragment near the well is
ⓐ. more negatively charged than the other fragment
ⓑ. stained with less ethidium bromide than the other fragment
ⓒ. smaller and therefore moved farther
ⓓ. larger and therefore moved a shorter distance through the gel
Correct Answer: larger and therefore moved a shorter distance through the gel
Explanation: Agarose gel acts as a molecular sieve, so DNA fragments are separated mainly on the basis of size. Larger fragments move more slowly and therefore remain closer to the loading well. Smaller fragments pass more easily through the gel matrix and travel farther. The difference is not mainly due to charge because DNA fragments carry a similar negative charge per unit length.
219. A plasmid contains a selectable marker and a suitable cloning site but lacks ori. What is the main consequence?
ⓐ. The plasmid will not replicate properly inside the host cell.
ⓑ. The plasmid will stain more strongly in UV light.
ⓒ. The inserted DNA will be expressed without a host.
ⓓ. The plasmid will cut itself at multiple sites.
Correct Answer: The plasmid will not replicate properly inside the host cell.
Explanation: The origin of replication is essential because it marks where DNA replication begins. Without ori, the plasmid cannot be maintained efficiently in the host even if it carries a selectable marker and a cloning site. As a result, the linked foreign DNA also fails to multiply properly. ori is therefore indispensable for plasmid persistence.
220. Which statement best distinguishes a selectable marker from a cloning site in a vector?
ⓐ. A selectable marker is where foreign DNA is inserted, whereas a cloning site controls plasmid copy number.
ⓑ. A selectable marker helps identify transformants, whereas a cloning site provides a defined position for DNA insertion.
ⓒ. A selectable marker initiates replication, whereas a cloning site protects the host DNA by methylation.
ⓓ. A selectable marker separates DNA fragments, whereas a cloning site causes colorless colonies.
Correct Answer: A selectable marker helps identify transformants, whereas a cloning site provides a defined position for DNA insertion.
Explanation: These two features serve very different functions in vector design. A selectable marker helps identify cells that have taken up the vector, often by antibiotic resistance or another detectable trait. A cloning site is the location at which foreign DNA is inserted into the vector. Confusing these two roles leads to many avoidable mistakes in recombinant DNA questions.
