Use of transgenic animals in target validation MCQs With Answer

This blog provides a focused set of multiple-choice questions on the use of transgenic animals for target validation in drug discovery, tailored for M.Pharm students. It explains how genetic modification of model organisms — including knockouts, knock‑ins, conditional systems, humanized models, and reporter lines — helps establish causality between a molecular target and disease phenotype. Questions cover genome editing methods (CRISPR/Cas9, ES-cell targeting), experimental design (timing, tissue specificity, phenotyping, pharmacological rescue), translational considerations (ADME/Tox, species differences), and ethical/regulatory aspects. These MCQs emphasize deeper understanding of methodological strengths, limitations, and practical implications for preclinical target validation and candidate selection.

Q1. What is the primary purpose of using transgenic animals in target validation during drug discovery?

• To increase compound solubility and formulation stability
• To demonstrate causal contribution of a gene/protein to a disease phenotype
• To replace clinical trials for regulatory approval
• To mass‑produce therapeutic proteins for commercial use

Correct Answer: To demonstrate causal contribution of a gene/protein to a disease phenotype

Q2. Which statement best describes the scientific rationale for creating a gene knockout model for target validation?

• To overexpress a human protein to study gain‑of‑function effects
• To remove gene function and observe whether disease phenotypes are reduced or absent
• To track protein localization using fluorescent tags
• To transiently inhibit a target in cell culture

Correct Answer: To remove gene function and observe whether disease phenotypes are reduced or absent

Q3. What advantage does a conditional (tissue‑specific) knockout using Cre‑lox provide over a constitutive global knockout?

• Permanent activation of the gene in all tissues
• Ability to control the timing and tissue in which the gene is deleted
• Guaranteed elimination of compensatory mechanisms
• Faster generation of founders compared with CRISPR editing

Correct Answer: Ability to control the timing and tissue in which the gene is deleted

Q4. Why are inducible systems (e.g., tamoxifen‑dependent Cre‑ER or Tet‑On/Tet‑Off) important for target validation?

• They permanently integrate reporter genes into the genome
• They enable temporal control to separate developmental from adult functions
• They automatically humanize mouse genes
• They eliminate the need for genotyping founders

Correct Answer: They enable temporal control to separate developmental from adult functions

Q5. How do humanized transgenic mice improve ADME/Tox and pharmacokinetic predictions for human drug candidates?

• By expressing human metabolic enzymes or transporters to mimic human drug metabolism
• By always producing identical plasma half‑lives to humans
• By eliminating all species differences so translation is perfect
• By increasing animal lifespan for chronic studies

Correct Answer: By expressing human metabolic enzymes or transporters to mimic human drug metabolism

Q6. Compared with traditional embryonic stem (ES) cell homologous recombination, what is a major advantage of CRISPR/Cas9 for generating transgenic models?

• It guarantees zero off‑target mutations in all cases
• It allows rapid, efficient editing directly in zygotes and enables multiplex edits
• It is the only method compatible with rats
• It requires no validation or founder screening

Correct Answer: It allows rapid, efficient editing directly in zygotes and enables multiplex edits

Q7. What is a common biological limitation when interpreting phenotypes from gene knockout models for target validation?

• Knockouts always produce identical phenotypes across different strains
• Compensatory upregulation of related genes can mask the true role of the target
• Knockouts eliminate the need for pharmacological validation
• They always predict human clinical outcomes accurately

Correct Answer: Compensatory upregulation of related genes can mask the true role of the target

Q8. What is the key use of reporter transgenic lines (e.g., GFP or luciferase under a target promoter) in target validation?

• To permanently silence the target gene across the organism
• To visualize and quantify spatial and temporal expression dynamics of the target
• To produce therapeutic antibodies in vivo
• To replace pharmacokinetic assays in drug discovery

Correct Answer: To visualize and quantify spatial and temporal expression dynamics of the target

Q9. When generating a disease‑associated point mutation in a gene, why is a knock‑in model preferred over transgenic overexpression?

• Knock‑ins place the mutation in the native genomic context and preserve physiological regulation
• Transgenic overexpression always reproduces human disease faithfully
• Knock‑ins avoid the need to breed animals
• Transgenic overexpression cannot cause gain‑of‑function phenotypes

Correct Answer: Knock‑ins place the mutation in the native genomic context and preserve physiological regulation

Q10. Which strategy most directly reduces off‑target editing when using CRISPR/Cas9 to create a transgenic model?

• Using very short guide RNAs with low specificity
• Adopting high‑fidelity Cas variants and well‑validated, specific guide RNAs
• Injecting excess Cas9 protein to overwhelm the cell
• Avoiding sequencing of founders to speed up experiments

Correct Answer: Adopting high‑fidelity Cas variants and well‑validated, specific guide RNAs

Q11. For robust target validation, why is pharmacological rescue (reversing phenotype with a drug) used alongside genetic perturbation?

• It proves that the genetic alteration is irrelevant to the phenotype
• It demonstrates that the phenotype is due to target modulation and is pharmacologically tractable
• It eliminates the need for control animals
• It increases the complexity so results are less interpretable

Correct Answer: It demonstrates that the phenotype is due to target modulation and is pharmacologically tractable

Q12. Which rationale supports choosing rats instead of mice for certain transgenic target validation studies?

• Rats are always cheaper and faster to generate than mice
• Rats often provide superior behavioral, surgical, and PK/ADME readouts due to larger size and physiology
• Rats never require genotyping
• Mice cannot be genetically modified

Correct Answer: Rats often provide superior behavioral, surgical, and PK/ADME readouts due to larger size and physiology

Q13. What is the main benefit of using bacterial artificial chromosome (BAC) transgenics for reporter or humanization studies?

• BACs exclude regulatory regions for simpler expression
• BACs can carry large genomic fragments including regulatory elements to reproduce endogenous expression patterns
• BAC transgenics always avoid position‑effect variegation
• BACs reduce the need for breeding to homozygosity

Correct Answer: BACs can carry large genomic fragments including regulatory elements to reproduce endogenous expression patterns

Q14. What does germline mosaicism in genome‑edited founders mean for downstream breeding and validation?

• All offspring will always inherit the same edited allele
• Founders can produce offspring with different alleles; extensive genotyping of progeny is required
• Mosaicism guarantees homogenous phenotype across tissues
• Mosaic founders cannot transmit edits to offspring

Correct Answer: Founders can produce offspring with different alleles; extensive genotyping of progeny is required

Q15. In cancer target validation, what is a key advantage of genetically engineered mouse models (GEMMs) over human tumor xenografts?

• GEMMs lack an intact immune system
• GEMMs model tumor initiation and microenvironment in an immune‑competent host
• Xenografts always better predict immunotherapy outcomes
• GEMMs are faster and require no breeding

Correct Answer: GEMMs model tumor initiation and microenvironment in an immune‑competent host

Q16. What is the definition of a “floxed” allele in the context of conditional genetics?

• An allele containing flanking loxP sites that permit Cre‑mediated excision
• An allele permanently silenced by methylation
• An allele inserted into a bacterial artificial chromosome
• An allele that expresses a fluorescent reporter constitutively

Correct Answer: An allele containing flanking loxP sites that permit Cre‑mediated excision

Q17. How does the tamoxifen‑inducible Cre‑ER system control Cre activity?

• Tamoxifen degrades Cre recombinase to prevent recombination
• Tamoxifen binds the estrogen receptor fusion, enabling nuclear translocation of Cre and recombination
• Tamoxifen permanently activates all promoters in the genome
• Tamoxifen introduces loxP sites into the DNA

Correct Answer: Tamoxifen binds the estrogen receptor fusion, enabling nuclear translocation of Cre and recombination

Q18. In biomarker validation using transgenic models, what is a critical demonstration that supports translational relevance?

• Showing that biomarker levels change in animals but cannot be measured in humans
• Linking target modulation genetically and pharmacologically to changes in a biomarker that is measurable in patients
• Demonstrating that the biomarker is only present in embryonic tissues
• Showing that the biomarker changes only after lethal dosing

Correct Answer: Linking target modulation genetically and pharmacologically to changes in a biomarker that is measurable in patients

Q19. Which ethical and regulatory principle is most important when planning transgenic animal experiments for target validation?

• Avoiding oversight to accelerate research timelines
• Applying the 3Rs (Replacement, Reduction, Refinement) and obtaining appropriate ethical review and justification
• Using as many animals as possible for statistical power without refinement
• Prioritizing only cost reduction regardless of welfare

Correct Answer: Applying the 3Rs (Replacement, Reduction, Refinement) and obtaining appropriate ethical review and justification

Q20. Why is backcrossing a newly generated transgenic allele into a defined inbred background important for target validation studies?

• To increase genetic variability and complicate phenotype interpretation
• To place the allele on a consistent genetic background and minimize modifier effects that could confound phenotype interpretation
• To avoid the need for proper controls
• To make genotyping unnecessary in future generations

Correct Answer: To place the allele on a consistent genetic background and minimize modifier effects that could confound phenotype interpretation

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