PCR Basics
What is the main purpose of PCR?
To make many copies of a specific DNA segment so it can be studied or analyzed.
A PCR reaction produces very little DNA. Name one possible reason this might happen.
Possible reasons include missing primers, inactive polymerase, incorrect temperatures, or too little starting DNA.
What is the main purpose of gel electrophoresis?
To separate DNA fragments based on size so they can be compared.
A DNA fragment traveled a cm from the well is it: large or small?
Large
Why is PCR often performed before gel electrophoresis in forensic investigations?
PCR increases the amount of DNA so there is enough to visualize during electrophoresis.
Why is heat used at the beginning of each PCR cycle?
Heat separates (denatures) the two DNA strands so they can be copied.
If primers fail to bind during PCR, which step of the process will be directly affected? What step is in directly affected?
Annealing is most directly affected, because primers must bind during this step; without primer binding, extension cannot occur (DNA polymerase cannot add nucleotides).
Why do DNA fragments move toward the positive end of the gel?
DNA has a negative charge, so it is attracted to the positive electrode.
If two DNA samples show matching bands at the same positions, what does that suggest?
The DNA fragments are similar in size and may have come from the same source or be closely related.
How can gel electrophoresis help determine biological relationships?
By comparing banding patterns between individuals to see shared or matching fragments.
What role do primers play in the PCR process?
Primers bind to specific DNA sequences and provide a starting point for DNA polymerase to begin copying.
A scientist accidentally uses a polymerase that cannot tolerate high temperatures. Predict the outcome and explain why.
No amplification would occur because the polymerase would denature during the high-temperature steps of PCR.
How does fragment size affect how far DNA travels through the gel?
Smaller fragments move faster and farther through the gel than larger fragments.
A gel shows multiple bands in one lane. What does this indicate about the DNA sample?
The sample contains multiple DNA fragments of different sizes.
Why would DNA fingerprinting be unreliable if only one DNA region were analyzed?
One region may match by chance; multiple regions provide stronger evidence.
Why is a heat-tolerant Taq polymerase necessary for PCR?
PCR involves high temperatures; a heat-tolerant polymerase (like Taq) will not denature and can function during the process.
A PCR tube is missing one key ingredient. No DNA bands appear after electrophoresis. Which type of ingredient(s) is most likely missing, and why?
Primers or DNA polymerase are most likely missing; without them, DNA cannot be copied and no bands would appear on the gel.
Why is agarose gel useful for separating DNA fragments?
Agarose forms a porous matrix that allows smaller DNA fragments to move through more easily than larger ones.
Why is analyzing multiple DNA regions more reliable than analyzing just one region?
Comparing multiple regions reduces the chance of a coincidental match and increases reliability.
Can gel electrophoresis always definitively identify a suspect on its own? Why?
Gel electrophoresis shows fragment size similarities but does not prove identity without additional analysis.
Explain why PCR targets specific DNA regions instead of copying the entire genome.
Targeting specific regions allows scientists to amplify only useful or variable DNA, making analysis faster, more efficient, and more informative.
A sample is contaminated with non-target DNA. Explain how this could affect PCR results and the appearance of the gel.
Contamination could cause extra or unexpected bands, making it difficult to determine which DNA came from the intended sample.
Explain why two DNA samples from different people might produce different banding patterns even if the same genes are analyzed.
Individuals have differences in DNA fragment lengths, producing unique banding patterns.
Two samples have mostly matching bands but differ in one key location. What conclusions can and cannot be made from this evidence?
They could be related, they are not from the same individual.
Identical twins are suspects in a crime. Explain why standard DNA analysis may fail to distinguish between them and what type of evidence might be needed instead.
Identical twins have the same DNA, producing identical banding patterns; other evidence such as fingerprints, epigenetic markers, or non-DNA evidence would be needed.