Gene libraries and cDNA libraries MCQs With Answer

Introduction: This quiz collection on Gene libraries and cDNA libraries is tailored for M.Pharm students studying Advanced Pharmaceutical Biotechnology. It covers construction, types, vectors, cloning strategies, screening methods, and applications of genomic and cDNA libraries with emphasis on laboratory techniques and interpretation. Questions probe concepts such as library complexity, insert size selection, oligo-dT versus random priming, full-length cDNA recovery, RACE, normalization, subtractive libraries, and screening methods like colony and plaque hybridization. The MCQs are designed to test both theoretical understanding and practical considerations encountered in constructing and using gene libraries for drug discovery, functional genomics, and recombinant protein expression.

Q1. What is the primary difference between a genomic library and a cDNA library?

• Genomic library contains only expressed sequences while cDNA library contains entire genome including introns
• Genomic library contains entire genomic DNA including introns and regulatory regions while cDNA library represents only expressed mRNAs converted to DNA
• Genomic library is constructed using reverse transcriptase while cDNA library is constructed using restriction enzymes
• Genomic library can only be created from prokaryotes while cDNA libraries are only from eukaryotes

Correct Answer: Genomic library contains entire genomic DNA including introns and regulatory regions while cDNA library represents only expressed mRNAs converted to DNA

Q2. Which enzyme is essential for synthesizing first-strand cDNA from mRNA during cDNA library construction?

• DNA ligase
• Reverse transcriptase
• DNA polymerase I
• RNA polymerase II

Correct Answer: Reverse transcriptase

Q3. Oligo(dT) primers are used in cDNA synthesis to selectively prime which region of mRNA?

• 5′ cap structure
• Poly-A tail at the 3′ end
• Internal ribosomal entry sites (IRES)
• tRNA-like structures

Correct Answer: Poly-A tail at the 3′ end

Q4. Which vector type is most suitable for cloning very large genomic DNA fragments (hundreds of kb)?

• Plasmid vectors
• Lambda phage vectors
• Bacterial artificial chromosome (BAC) or yeast artificial chromosome (YAC)
• Single-stranded M13 vectors

Correct Answer: Bacterial artificial chromosome (BAC) or yeast artificial chromosome (YAC)

Q5. In library construction, “coverage” refers to:

• The percentage of clones that are recombinant
• The average number of times each base of the genome is represented in the library
• The number of different vectors used
• The fraction of mRNA converted to cDNA

Correct Answer: The average number of times each base of the genome is represented in the library

Q6. Which method is commonly used to screen a genomic or cDNA library for a specific sequence using a labeled nucleic acid probe?

• Colony or plaque hybridization
• Flow cytometry
• Western blotting
• Mass spectrometry

Correct Answer: Colony or plaque hybridization

Q7. Random priming during cDNA synthesis is used to:

• Preferentially synthesize full-length 5′ ends
• Generate cDNA from both polyadenylated and non-polyadenylated RNA sequences and increase coverage of 5′ regions
• Only transcribe rRNA
• Remove introns from genomic DNA

Correct Answer: Generate cDNA from both polyadenylated and non-polyadenylated RNA sequences and increase coverage of 5′ regions

Q8. What is the purpose of linkers or adaptors in library construction?

• To degrade mRNA before cloning
• To provide restriction sites and facilitate ligation of inserts into cloning vectors
• To fluorescently label DNA for sequencing
• To prevent bacterial transformation

Correct Answer: To provide restriction sites and facilitate ligation of inserts into cloning vectors

Q9. A normalized cDNA library is designed to:

• Increase the representation of highly abundant transcripts
• Reduce redundancy and equalize representation of rare and abundant transcripts
• Eliminate all rare transcripts
• Only contain full-length transcripts

Correct Answer: Reduce redundancy and equalize representation of rare and abundant transcripts

Q10. Subtractive hybridization is mainly used to:

• Construct genomic libraries from fragmented DNA
• Enrich for sequences unique to one sample by removing sequences common to both samples
• Sequence entire genomes rapidly
• Clone vectors without inserts

Correct Answer: Enrich for sequences unique to one sample by removing sequences common to both samples

Q11. Which technique helps obtain full-length cDNA clones when 5′ ends are underrepresented?

• RACE (Rapid Amplification of cDNA Ends)
• Southern blotting
• Restriction digestion with blunt cutters
• DNA footprinting

Correct Answer: RACE (Rapid Amplification of cDNA Ends)

Q12. Which statement about expression libraries is correct?

• They are constructed so that cloned inserts are not transcribed or translated
• They use vectors that allow transcription and translation of cloned inserts to screen for protein function or antigenicity
• They only contain genomic DNA fragments with introns
• They cannot be used to identify antigens recognized by antibodies

Correct Answer: They use vectors that allow transcription and translation of cloned inserts to screen for protein function or antigenicity

Q13. Which screening method is most appropriate for identifying clones expressing a protein that binds a specific ligand?

• Colony PCR
• Immunoscreening or ligand-binding assays on expression libraries
• ELISA of genomic DNA
• Restriction fragment length polymorphism (RFLP)

Correct Answer: Immunoscreening or ligand-binding assays on expression libraries

Q14. Why are lambda phage vectors historically popular for genomic libraries?

• They can package and efficiently clone intermediate-sized inserts (~15–20 kb) and form plaques for easy screening
• They can replicate in eukaryotic nuclei
• They integrate stably into the bacterial chromosome
• They exclusively clone very large inserts (>200 kb)

Correct Answer: They can package and efficiently clone intermediate-sized inserts (~15–20 kb) and form plaques for easy screening

Q15. In calculating the number of clones required for a genomic library with desired probability P of containing a given sequence, which parameter is directly used?

• Insert GC content
• Genome size and average insert size
• Vector antibiotic resistance marker
• Number of restriction sites

Correct Answer: Genome size and average insert size

Q16. ESTs (Expressed Sequence Tags) are generated from:

• Short genomic DNA fragments to identify introns
• Single-pass sequencing of cDNA clones to provide partial sequences of expressed genes
• Sequencing of plasmid backbones only
• Proteomic mass spectra converted to DNA sequences

Correct Answer: Single-pass sequencing of cDNA clones to provide partial sequences of expressed genes

Q17. When constructing a cDNA library intended for heterologous expression in bacteria, which strategy improves the chance of obtaining functional protein expression?

• Cloning random genomic fragments including introns
• Using cDNA synthesized from mRNA, with selection for full-length ORFs and insertion into an expression vector with bacterial promoter and ribosome-binding site
• Using only ribosomal RNA as input
• Cloning exclusively mitochondrial DNA

Correct Answer: Using cDNA synthesized from mRNA, with selection for full-length ORFs and insertion into an expression vector with bacterial promoter and ribosome-binding site

Q18. Which of the following is a disadvantage of cDNA libraries compared to genomic libraries?

• cDNA libraries include intronic and regulatory sequences
• cDNA libraries do not represent non-expressed genes or regulatory regions and often miss 5′ ends of transcripts
• cDNA libraries always have too large inserts for plasmid vectors
• cDNA libraries cannot be used to express proteins

Correct Answer: cDNA libraries do not represent non-expressed genes or regulatory regions and often miss 5′ ends of transcripts

Q19. Which factor most directly reduces redundancy when creating a normalized cDNA library?

• Using only oligo(dT) priming
• Equalizing transcript frequencies by hybridization-based depletion or reassociation kinetics to remove abundant cDNAs
• Using longer restriction enzymes
• Increasing vector copy number

Correct Answer: Equalizing transcript frequencies by hybridization-based depletion or reassociation kinetics to remove abundant cDNAs

Q20. In plaque hybridization of a lambda phage genomic library, what is typically transferred to the membrane for probing?

• Intact phage particles only
• Bacterial chromosomal DNA
• Nucleic acids from phage plaques (DNA) bound to nitrocellulose or nylon membrane
• Protein lysates of host cells

Correct Answer: Nucleic acids from phage plaques (DNA) bound to nitrocellulose or nylon membrane

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