Laboratory Techniques Practice Questions

Correct answer: Amplify a specific DNA sequence

PCR (Polymerase Chain Reaction) is used to make millions of copies of a specific DNA segment in vitro. This amplification allows detailed analysis even from very small or degraded DNA samples.

Correct answer: Taq DNA polymerase

Taq DNA polymerase is a thermostable enzyme that synthesizes DNA at high temperatures. It is crucial for PCR because it remains active after repeated heating cycles.

Correct answer: Denaturation

Denaturation involves heating the DNA (typically to ~95 °C) to break the hydrogen bonds between complementary strands. This produces single-stranded DNA templates available for primer binding and amplification.

Correct answer: Primer annealing

Annealing occurs at a lower temperature (typically 50–65 °C), allowing primers to bind to their complementary sequences on the template DNA. This step determines the specificity of PCR amplification.

Correct answer: Charge and size

Gel electrophoresis separates molecules by size and charge as they migrate through a gel under an electric field. Smaller molecules generally move faster through the gel matrix.

Correct answer: RNA and DNA fragments

Agarose gels are widely used to separate nucleic acids such as DNA and RNA based on size. The porous gel matrix allows fragments to migrate at different rates through an electric field, enabling efficient size-based separation of nucleic acid mixtures.

Correct answer: Positive electrode

DNA has a negatively charged phosphate backbone, so it moves toward the positive electrode during electrophoresis. This property enables its separation in an electric field.

Correct answer: Ethidium bromide

Ethidium bromide intercalates between DNA base pairs and fluoresces orange under UV light, making DNA bands visible after gel electrophoresis. It is one of the most widely used DNA stains, though safer alternatives such as SYBR Safe are now also common.

Correct answer: Estimate fragment size

A DNA ladder contains fragments of known sizes that serve as molecular weight markers. Comparing sample bands with the ladder helps estimate fragment lengths.

Correct answer: Primer sequences

Primer sequences are short, single-stranded DNA oligonucleotides that bind to specific complementary target regions on the template. They define the start and end points of the amplification region and provide the free 3'-OH group that DNA polymerase requires to begin synthesis.

Correct answer: Separation of mixtures

Chromatography separates the components of a mixture based on their differential movement between a stationary phase and a mobile phase. It is widely used in biochemical analysis, pharmaceutical purification, and quality control of compounds.

Correct answer: Filter paper

In paper chromatography, the paper acts as the stationary phase. The solvent moves through the paper carrying the components at different rates.

Correct answer: The carrier solvent system

The mobile phase is the solvent or gas that carries the sample mixture through the stationary phase. Because different components have different affinities for the stationary and mobile phases, they travel at different speeds and are thereby separated.

Correct answer: Paper chromatography

Paper chromatography is often used to separate plant pigments based on their solubility and movement with the solvent. It is a simple and effective analytical method.

Correct answer: Relative flow of solute

The Rf (retardation factor) value is the ratio of the distance traveled by the solute to the distance traveled by the solvent front under the same conditions. It is a characteristic constant for a given substance in a specific solvent system and is used for compound identification.

Correct answer: Gel filtration chromatography

Gel filtration chromatography (size-exclusion chromatography) separates molecules based on size as they pass through porous beads. Larger molecules cannot enter the pores and elute first, while smaller molecules enter the pores and take longer to pass through.

Correct answer: Repeated thermal cycling steps

PCR requires repeated cycles of denaturation (high heat to separate strands), annealing (cooling for primer binding), and extension (moderate heat for polymerase synthesis). These thermal cycles enable exponential amplification of the target DNA sequence.

Correct answer: Proteins and small DNA fragments

Polyacrylamide gels have smaller, more uniform pore sizes than agarose gels, allowing high-resolution separation of proteins and small nucleic acid fragments. They are routinely used in SDS-PAGE for protein analysis and in sequencing applications.

Correct answer: PCR amplification

PCR (Polymerase Chain Reaction) selectively amplifies a specific gene sequence, making it detectable even from minute initial quantities. This selectivity makes PCR the gold standard for gene identification, clinical diagnostics, and forensic analysis.

Correct answer: All are molecular analysis and separation tools

PCR amplifies specific DNA sequences, gel electrophoresis separates biomolecules by size and charge, and chromatography fractionates chemical mixtures by differential affinity. Together, these techniques are core analytical tools across all branches of modern molecular biology and biochemistry.