Download Lab 11: DNA Testing

Name________________
Section_______________
Spring 2007
DNA Testing Lab
Biology 212
Genetics Laboratory
Goals of the Lab: To expose students to technologies used for DNA testing and
applications for their use in medical diagnostics and forensics.
Reference and Background Information: European Initiative for Biotechnology
Education at http://www.eibe.info/
Lab Procedure for DNA Testing Simulation
1. A 0.6% (w/v) agarose gel in 1xTBE must be prepared about a half hour ahead
of the lab period. Most likely, we will prepare gels the day before lab and store
them for use the next day. For the small gels, weigh out 0.3 g agarose, add 50 ml
1xTBE and heat for about 3 min. in the microwave. Add 2 microliters of 1 mg/ml
ethidium bromide after the solution has cooled slightly. Pour into a gel tray that
has been sealed on the ends with tape. A comb should be placed near one end.
Larger gels require 0.6 g agarose/100 ml 1xTBE. The gel should set for about
20-30 min. before use.
2. To set up the gels, put on gloves (ethidium bromide is a carcinogen!), and
remove the tape from the ends of the gel. Place in the gel apparatus and cover
the gel with 1xTBE electrophoresis buffer. Make sure the tray is oriented so that
the wells are located closer to the black (negative) pole and away from the red
(positive) pole.
3. Loading DNA samples:
a. Each group of 2 or 3 should load a set of four samples. If you have two
groups sharing a gel, you should run duplicate sets of samples.
b. The samples have been prepared for you and contain the DNA sample mixed
with loading buffer. Load 10 μl of each sample/well.
Lane
1
2
3
4
Sample
Suspect 1
Suspect 2
Suspect 3
Evidence
4. After the group’s samples are loaded, connect the cover and the electrodes
so that the DNA samples will migrate toward the positive (red) pole. Insert the
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red and black electrodes into the power supply with the power turned off. Turn
the power on and adjust the voltage to 100 volts. Carry out the electrophoresis
for about 1 hour or until the blue dye front has migrated one half to two thirds the
distance down the gel.
5. Your gel will be photographed using the imaging workstation.
 Be sure to provide the instructor with information about your gel and which
students worked together.
 You will be provided with a color printout; images can also be emailed at
your request. Image files are rather large, so you might find it easiest to
open a word file, insert the gel file as a picture, then resize the picture to
approximately its normal size.
Worksheet DNA Testing Lab (20 points)
Worksheet due Wed. May 9, 2007 in class.
Analysis of results:
1. Provide a picture of the gel electrophoresis result you obtained in lab. Label
the lanes and attach to this assignment.
2. Based on the results, which suspect’s DNA pattern matchs that of the
evidence at the crime scene?
3. a. Prepare a table of standard and unknown fragments. The fragment sizes
from lane 1 (Suspect 1) are given. Measure the distance each standard
fragment (in cm or mm) migrated using a centimeter ruler. Use this to construct a
standard curve on semi-log graph paper. Be sure to label the axes of the graph.
Lane 1:Suspect 1
Lane 2:Suspect 2 Lane 3:Suspect 3 Lane 4:Evidence
distance
base
distance base distance base
distance
base
pairs
pairs
pairs
pairs
23,000
9400
6000
4400
(faint)
2200
2000
560
125
(usually
not
seen)
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b. Describe some specific differences in the DNA banding pattern of the
suspects whose DNA does not match the DNA pattern of the evidence. For full
credit, use particular band sizes in your explanation.
4. Compare the DNA patterns of suspect 1 and suspect 2.
a. Determine the sizes of any fragments that significantly differ between these
two individuals, using the size information from question 3.
b. How related are suspect 1 and suspect 2 likely to be, based on their DNA
patterns?
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5. a. Select one of the following methods or applications or another technology
relevant to the study of human genetics or population genetics and briefly
research it, using the web site of the European Initiative for Biotechnology
Education, your textbook, and/or any other resources you choose.
Topics:
 RFLP analysis
 PCR-based DNA profiling
 PCR-based DNA tests
 RAPD analysis
 SNP analysis
 Microarrays (DNA chips)
 Positional cloning
 Pyrosequencing
 Real-time PCR
 Reverse Transcriptase PCR
 RNA interference
 Transgenic mice
 Fluorescent in situ hybridization (FISH)
 Allele specific oligonucleotide hybridization
 Northern blot
 Spectral karyotype analysis (SKY)
 Taq-Man assay
b. In an answer of one to two pages at most, in your own words define or briefly
explain what the method or application entails, describe how it might be carried
out, and provide a specific example of what you might learn from it or how it
might be applied (historical examples are fine). Your answer should be neatly
handwritten or computer printed, double-spaced, and enclosed with your
worksheet. Be sure to list the references you consulted. For web sites, give the
URL. For text or journal sources, give author, title, year, volume (journal), and
pages.
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