A: Background Information
DNA fingerprinting is used in many important cases around the world. One of the most important and popular one that is very well known is in crime scene investigations. Human DNA samples will be taken from the crime scene are are compared to different suspects and DNA fingerprints will help figure out who really committed the crime. Although TV shows make it look easy, this lab will help students find out that is much more complicated it really is. Here is some background information you will need to know before this lab. The DNA samples for different "suspects" will be provided however, there is more you have to do with the samples, like with restriction enzymes. Restriction enzymes make cuts at specific sequence of base pairs at a palindrome and comes from bacteria. In addition, restriction enzymes can occur in different locations multiple times, resulting in many different fragments. Therefore, in this lab we will use the restriction enzyme EcoRL to cut strands of circular plasmid DNA in the samples provided for us. This cut DNA can be seperated and observed using a process called agarose gel electrophoresis, also known as RFLP (Restriction fragment length polymorphism).
In agarose gel electrophoresis, DNA fragments are separated by size. One will notice this when the DNA samples, enzyme, buffer and dye is added into the agarose gel. Inside the gel there is a chamber for each different sample, and once added into the chamber time is needed to create these different sizes. During that time, a direct current is passes through electrodes at each end of the chamber. Since the DNA is negatively charged, it will be attracted to the positive pole. It is because of this that when the process is done, one sees bands of DNA. These strands are known as DNA fingerprints.
Each person has their own DNA fingerprint, just like a real fingerprint. It is the DNA's radioactive probes which recognize and bind a sequence to create a "radioactive tag" that will produce different blotting bands. The size and length of these bands reflect variations in the individuals' DNA. These bands can be seen by adding a loading dye to the DNA samples before injecting them into the agarose gel. This loading dye will create a color that will allow you to see the different types of bands, in addition to making the sample heavier so it will sink to the bottom of the solution. This DNA can be taken from any biological material that contains DNA, such as: body tissues, bodily fluids (blood and semen), hair follicles, ect and can be used even if this material is dried.
B: Objective & Purpose
The objective and purpose of this lab is to use the DNA provided to figure out who committed the crime through the different instructions above and below. With all the different DNA samples, enzymes, loading dye and agarose gel, one will be able to figure out who the real "killer" is. However, DNA fingerprinting is not only used in crime scenes but also for other cases, such as: food identification, to set accused and convicted felons free, identifying human remains, determining relatedness of humans, studying relatedness among ancient peoples, DNA testing of families, identifying organisms that cause disease, identifying birth parents (paternity testing), proving paternity, determining effectiveness of bone marrow transplants, proving relatedness of immigrants, confirming relatedness among animals and finally, DNA testing of plant materials. So from, plants to immigrants to convicts, DNA fingerprinting is used everywhere to improve and excel our scientific studies.
C: Procedures
In the Forensic DNA Fingerprinting Lab, the DNA from the "crime scene" will be extracted so that it can be tested with "suspect's" DNA to figure out who did the crime! On the first day (Lesson 1 - Restriction Digestion), one will first label six empty tubes with Suspects 1-5 (label S_ [S1 in blue, S2 in orange, S3 in violet, S4 in red, S5 is yellow]) and the one remaining Crime Scene into the green tube, in addition to putting your period and lab table on the micro test tube holder. Afterward, each lab partner will take one or two of those empty tubes to the front of the class and fill them up with the corresponding suspects or crime scenes DNA. Once that is done, a lab partner will take the enzyme mix from the ice container and add it all the DNA samples while mixing them with the pipets. Then with the microcentrifuge available at one's lab table, spin the centrifuge to collect all the liquid from the bottom of the tube. The last step in Lesson 1 is to place the tubes in the foam micro tube holder and incubate overnight at 37 degrees C.
Lesson 2: Agarose Gel Electrophoresis will proceed day one (Lesson 1). First a lab partner will get the DNA samples from incubator and put them all into a centrifuge and spin it for only five seconds. Then using a separate tip for each sample, so that the samples don't get contaminated, add the DNA loading dye to all the samples. This DNA loading dye will show us the DNA and makes the sample heavier. If it is not already done so, place the agarsose gel provided in the electrophoresis apparatus and fill the chamber with TAE butter to cover the gel. Then check that the wells of the agarose gels are near the black (-) electrode and the bottom red (+) electrode. Afterward, use a separate tip and load each sample into the 7 wells of the gel in the following order: Lane 1 - DNA size marker (just dye); 10 ul, Lane 2 - Crime Scene; 20 ul, Lane 3-7 - S1-S5; 20 ul. Once done, carefully place the lid of the electrophoresis chamber and connect the Red and black jacks on the lid of the horizontal electrophoresis chambers so that they match with the red and black jacks on the base and plus in the red to red and balck to black on the power supply. Make sure the power and electrophorese is at 200 V for 12 minutes. Wait 24 hours and the samples will be show who is "guilty of the crime"!
D: Controls, Variables & Prediction
The variables in this lab are going to be all the different suspects which we have DNA samples for. These are the variables for this lab because they are all different and we will use them to find out who committed the crime. However, the control of the lab will be the DNA found at the crime scene. This is the control because it will stay the same and the other DNA samples will have to match it. In this lab we are unable to make a prediction on what will happen because it is necessary to get the DNA to predict who committed the crime.
Observations:
DNA Evidence!
Discussion:
A: Analyze and Explain Data
The data that we received from this lab proved that suspect 3, Katie Records, "committed this murder". We found this out by comparing the DNA evidence found at the "crime scene" to the DNA sample of Miss. Records. In the agarose gel, the DNA samples matched, it was hard evidence that she had committed this crime.
B: Possible Sources of Error
Some possible sources of error in this lab are if one did not change the pipets every time they went between DNA samples which would contaminate the samples. In addition, one could have also mixed the test tubes or mixed the samples while putting them into the agarose gel and that would completely throw off ones results causing them to receive the wrong information. However, if one pays attention to the lab, is careful and avoided sources of error; the lab will go perfectly and one will find out who the criminal is!
No comments:
Post a Comment