WET LAB DNA Barcoding: From Samples to Sequences - Northwest ...

WET LAB
KEY
Lab 4: Preparation of PCR Samples for
DNA Sequencing Teacher Answer Key
On your separate sheet of paper or in your lab notebook, answer each of the following questions:
1. Which band in your molecular weight standard is closest in size to your PCR product, and how much DNA
is in that molecular weight standard band?
Students should use either the 500 bp band or the 750 bp band. Both contain 25 ng of DNA.
2. Is your PCR product band brighter than the band in the molecular weight standard, as bright, or less bright?
Student answers will vary by experiment.
3. Based on your answers to Questions #1 and #2, how much DNA would you estimate is present in 10 μl of your
PCR product? (Remember, you added 10 μl of your PCR product to the agarose gel.)
Student answers will vary by experiment, but should be consistent with answers to Questions #1 and #2.
For example, if the student’s PCR product is half as bright as the 500 bp band containing 25 ng of DNA,
the student should estimate that their DNA band contains approximately 12.5 ng of DNA.
4. What is the estimated concentration of your PCR product (in ng/μl)? Show your work.
Student answers will vary by experiment, but should be consistent with answers to Questions #1–3.
For example, if the student estimated that their DNA band contained approximately 12.5 ng of DNA
(from Question #3):
12.5 ng DNA / 10 μl of PCR product loaded onto the gel = 1.25 ng/μl or 12.5 μg/ml.
5. What did you do in this laboratory experiment and why?
Purified the PCR product away from the PCR reaction components like primers, buffer, and DNA polymerase;
estimated the concentration of the PCR product.
6. What skills did you learn or practice?
This question is designed to help students identify laboratory skills that they can list on a resume and/or
college application.
• Handling samples
• Pipetting
• PCR product purification
Wet Lab – DNA Barcoding: From Samples to Sequences
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©Northwest Association for Biomedical Research—Updated October 2012