WorksheetsPCR and Electrophoresis
Total questions: 43
Worksheet time: 22mins
Which statement best defines a Single Nucleotide Polymorphism (SNP)?
A mutation replacing one nucleotide with another
A mutation deleting five consecutive nucleotides
A recombination swapping large chromosomal arms
A duplication creating an extra gene copy
Why can some SNPs change a protein without changing its amino acid sequence?
Frameshifts always preserve amino acids
Introns always encode amino acids
Codon degeneracy allows silent substitutions
Post-translational edits remove all SNP effects
Which restriction enzyme makes asymmetric sticky ends, and which makes blunt ends?
EcoRI sticky ends, HaeIII blunt ends
HaeIII sticky ends, EcoRI blunt ends
Both EcoRI and HaeIII sticky ends
Both EcoRI and HaeIII blunt ends
Which laboratory technique amplifies the TAS2R38 region to visualize enzyme effects?
CRISPR-Cas9 genome editing
Polymerase chain reaction (PCR) amplification
Southern blot with DNA probes
Sanger sequencing with chain terminators
What is the primary role of primers in PCR of TAS2R38?
Methylate cytosines in recognition sites
Ligate DNA fragments post-amplification
Proofread mismatched base pairs
Bind to template DNA to start synthesis
Which polymerase is commonly used in PCR due to heat stability?
Reverse transcriptase from retroviruses
Pfu polymerase from Pyrococcus furiosus
DNA pol I from Escherichia coli
Taq polymerase from Thermus aquaticus
During PCR, which step involves primers binding to their complementary sequences on single-stranded DNA?
Extension by polymerase
Annealing at moderate temperature
Restriction enzyme digestion
Denaturation at high temperature
Which enzyme is responsible for synthesizing new DNA strands during the extension step of PCR?
Reverse transcriptase
RNase A nuclease
DNA ligase from E. coli
Taq DNA polymerase
What temperature is typically used for denaturation in PCR cycles?
25°C room temperature
94°C high heat
40–65°C range
72°C intermediate
Which statement best explains why PCR product doubles each cycle under ideal conditions?
Restriction enzymes cut and replicate DNA
New primers are synthesized de novo
Electrophoresis concentrates DNA molecules
Each template yields two complementary copies
In the PCR cycle diagram, what happens immediately after denaturation?
Restriction sites are methylated
Taq ligates DNA ends
Primers anneal to target sites
Fragments are separated by gel
Why is Taq DNA polymerase suitable for PCR?
It digests RNA primers during PCR
It creates blunt ends on DNA
It remains active at high temperatures
It binds sticky ends strongly
Restriction enzymes can produce sticky or blunt ends. Which describes sticky ends?
Symmetric straight cuts
Overhanging single-stranded ends
Circularized DNA products
No cleavage of phosphate backbone
During RFLP analysis, what directly causes differing fragment sizes among genotypes?
Different polymerase fidelity
Different primer melting temperatures
Presence or absence of restriction site
Variation in gel voltage
If the annealing temperature is set too high, which outcome is most likely?
Primers fail to bind efficiently
Non-specific primer binding increases
Taq denatures completely
Restriction sites are removed
Which step supplies dATP, dCTP, dGTP, and dTTP to build the complementary strand?
Denaturation separates strands
Electrophoresis sizes fragments
Annealing positions primers
Extension incorporates nucleotides
Which lab safety practice best minimizes exposure to hot reagents like boiling water and melted agarose?
Use longer microcentrifuge tubes
Stand farther from the water bath
Keep benches free of paper towels
Wear gloves and goggles consistently
During cheek cell collection, what is the primary role of rinsing vigorously with saline?
Dislodge epithelial cells into solution
Sterilize the oral cavity completely
Neutralize lysis buffer pH levels
Precipitate DNA directly from saliva
After the first centrifugation of the cheek cell suspension, which component should be poured off while leaving the desired material behind?
Supernatant is removed, pellet remains
Both pellet and supernatant are discarded
Pellet is removed, supernatant remains
Supernatant and pellet are mixed together
Which temperature and duration are used for the initial PCR denaturation step in the program provided?
55°C for 4 minutes
64°C for 30 minutes
94°C for 4 minutes
72°C for 5 minutes
Why is the lysis buffer added to the resuspended cheek cell pellet before incubation at 55°C and 99°C?
To burst cells and release DNA
To precipitate proteins with salt
To stain DNA for visualization
To cool the sample rapidly
Which step ensures that clumps of cells are not left in the sample after adding lysis buffer?
Lower the incubation temperature
Add more saline to dilute
Let the tube sit undisturbed
Vortex or flick the tube vigorously
What is the correct action after boiling the lysate at 99°C for 5 minutes?
Add primers for PCR directly
Pour off the pellet immediately
Centrifuge at full speed for 2 minutes
Store at room temperature overnight
Which statement best describes where the DNA is located after the second centrifugation of the lysate?
In the supernatant above cell debris
In the pellet with cell debris
Bound to the tube walls only
In the dense protein fraction
Which instruction prevents loss of the target material during the first pellet wash steps?
Invert the tube repeatedly
Do not disturb the cell pellet
Discard the pellet to remove salts
Add lysis buffer before pouring
What is the correct sequence for thermal cycling in the provided PCR program after initial denaturation?
99°C 5 min, 55°C 5 min, 72°C 5 min
94°C 30 s, 64°C 30 s, 72°C 30 s
72°C 5 min, 55°C 5 min, 99°C 5 min
64°C 30 s, 72°C 30 s, 94°C 4 min
Why is labeling microcentrifuge tubes at the start of Module I critical for experiment workflow?
Enhances DNA yield through tracking
Speeds up boiling during incubation
Improves centrifuge rotor balance
Prevents sample mix-ups between groups
Which combination correctly represents the core PCR reaction mix for amplifying the PTC region?
30 µL primer mix, 5 µL DNA, dNTPs only
10 µL primer mix, 40 µL DNA, Taq polymerase only
40 µL primer mix, 10 µL DNA, EdvoBead PLUS
20 µL primer mix, 25 µL DNA, MgCl2 buffer
During PCR thermal cycling, what is the correct order of steps within each cycle?
Annealing, extension, denaturation
Extension, denaturation, annealing
Denaturation, annealing, extension
Denaturation, extension, annealing
A student mixes the PCR sample and sees undissolved EdvoBead PLUS. What is the best immediate action?
Continue mixing gently until it dissolves
Increase primer concentration drastically
Skip mixing and start thermal cycling
Add extra extracted DNA to dilute beads
Which thermal cycling condition is most likely the annealing temperature used in this protocol?
72°C for 5 minutes
72°C for 30 seconds
65°C for 30 seconds
94°C for 30 seconds
Why is an undigested PCR sample saved when setting up the HaeIII restriction digest?
It prevents evaporation during incubation
It serves as a control for comparison on the gel
It increases enzyme activity during digestion
It provides a higher primer concentration
In this protocol, which step ensures the sample is collected at the bottom of the tube before incubation?
Storing at -20°C first
Adding more primer mix
Heating at 94°C briefly
Brief centrifugation after mixing
What sequence feature does HaeIII recognize and cut when present in the PCR product?
Random nucleotide repeats
Only single-stranded DNA
AT-rich promoter regions
GGCC sequence sites
Which item should be loaded first according to typical lane order?
Control digested in lane 6
Student undigested in lane 1
DNA ladder in lane 1
Empty lane reserved in lane 3
What is the correct procedure after electrophoresis completes?
Remove gel and tray from chamber
Increase voltage to sharpen bands
Add fresh buffer to cool the gel
Flip polarity and run five more minutes
How should the gel be positioned for visualization on a transilluminator?
Invert the gel and remove the tray
Place the chamber directly on the glass
Slide the tray onto the viewing surface
Hold the gel in the air above light
Which safety practice is recommended when using a UV transilluminator?
Increase brightness to maximum
Remove buffer with bare hands
Wear UV-protective goggles
Look closely without shielding
Which statement correctly relates genotype to phenotype for PTC tasting?
TT weak taster, Tt non‑taster, tt strong taster
TT and tt both strong tasters, Tt non‑taster
TT often strong taster, Tt variable taster, tt non‑taster
TT non‑taster, Tt strong taster, tt weak taster
Which description best matches the three main PCR steps in a single cycle?
Denaturation freezes enzymes, annealing digests templates, extension removes nucleotides
Extension splits strands, denaturation builds primers, annealing elongates bases
Annealing melts DNA, extension cools reaction, denaturation copies primers
Denaturation separates strands, annealing binds primers, extension synthesizes DNA
If a DNA digest for TAS2R38 shows two distinct bands for one sample and a single band for another, what is the most likely interpretation?
Two bands mean non‑taster; one band means strong taster
Two bands indicate contamination; one band indicates purity
Two bands indicate heterozygote; one band indicates homozygote
Two bands mean failed PCR; one band means successful PCR
Which reasoning best explains why classmates with the same genotype might report different PTC tasting intensities?
PTC paper is chemically identical to control strips
Taste receptors never bind PTC in any genotype
PCR always mislabels genotypes in classroom labs
Environmental factors and perception variation affect phenotype
Which step ensures reliable comparison between control and PTC paper during tasting?
Share notes only after class discussion ends
Record observations for both strips immediately after tasting
Skip the control to avoid adaptation effects
Taste PTC multiple times without notes
