Solving the Mystery using Restriction Enzymes and Gel Electrophoresis

Introduction:
a) Restriction enzymes are a specific type of protein which can be found in bacteria. They act as a defense against viruses by cutting up viral DNA at specific locations called palindromes (these specific sites can also be called restriction sites). Gel electrophoresis is a process used to analyze DNA - the DNA is placed in a gel and an electric current is sent through. DNA is negatively charged, thus it will be pulled toward the positive node. However, the smaller, lighter pieces of DNA can move faster and travel farther along the gel. If this cut-up DNA is run on a gel, we can determine the length of each fragment of DNA based on how far they traveled along the gel. The different fragments can be seen as dark bands along the gel. This is called Restriction Fragment Length Polymorphism (RFLP).
b) The purpose of this experiment is to analyze different suspects' DNA via gel electrophoresis and compare their DNA to the crime scene's. The suspect whose DNA matches the crime scene's (meaning the bands are the same) is the culprit.
c) The purpose of the restriction enzymes is to cut up the suspects' DNA - each person has a unique genetic code, thus each person's DNA will be cut at different locations and will result in unique DNA fragment lengths. A loading buffer containing blue dyes is added to the DNA samples before injection into the gel - this allows us to monitor the process of gel electrophoresis. The gel electrophoresis and RFLP will then show a unique banding pattern of each person's DNA fragment lengths on a gel.  We can then analyze these bands to determine which DNA matches the DNA of the crime scene.
d) I cannot predict which person will be the culprit since I have not seen the results yet. However, the control is the crime scene DNA and the variables are the suspects' DNA.

Procedure:
Basically explained above...

Results:
It turns out Suspect #3 was the criminal! The DNA bands from Suspect #3 clearly matched the DNA bands of the Crime Scene - the evidence is incriminating.

Discussion:
a) Via gel electrophoresis, the fragments were separated by length along the gel - the shorter fragments traveled farthest toward the positive node while the longer fragments remained further behind. By comparing the bands, it was clear that the DNA bands of Suspect #3 exactly matched the bands of the Crime Scene.
b) Possible sources of error:

• Improper pipetting
• Contamination of DNA
• Not run on the gel long enough to accurately disperse DNA fragments

Creating biofuels from cellulose using enzyme cellobiase

Introduction:
a) Enzymes speed up the rate of chemical reactions without being used up by first binding to the reactants (substrate). The location of the binding is called the enzyme's active site. This bond lowers the activation energy (the energy needed to start the reaction), making the reaction occur faster. Enzymes' effectiveness can be affected by pH, temperature, and salinity. The reaction tends to occur faster if the concentration of enzymes or substrates is increased, but there is a point when the solution becomes saturated with either enzymes or substrates and cannot work any faster. The enzyme we will be using on this lab is cellobiase, which breaks down cellobiose - a sugar derivative of cellulose, a polysaccharide found in plant cell walls. The biofuel agency uses enzymes such as cellobiase to break down cellulose in plant matter and convert it to glucose. Glucose can then be converted to ethanol by microbial fermentation. Ethanol can then be used as an energy source to power certain motors and engines.
b) The purpose of this lab is to experience first hand in the classroom what biofuel companies do on a massive scale. Biofuel companies are constantly experimenting to find the most efficient way to make biofuels - and the method we will use in this experiment is only one method out of many.
c) We will use pipetting techniques to carefully and accurately transfer enzymes, buffers, and substrates to the solution. On Day #1, we will transfer a little bit of the solution at various time points to another test tube with a strong base to stop the reaction. The base will also turn p-nitrophenol (a product of the reaction) yellow, so we can judge how much product (including the glucose) has been created. On Day #2, we will have to carefully grind up mushrooms and add the mushroom extract to the reaction and measure how this affects the speed of the reaction.
d) I predict that, as time goes on, more product will be produced from the reaction. Also, I think the mushroom, which contains more enzymes, including cellobiase, will speed up the reaction rate. The variable of this experiment is the mushroom. The control is the amount of product produced under normal circumstances (Day #1).

Results/Observations:
The solution from Day #1 turned more yellow over time (the first cuvette was the lightest and the last cuvette, the darkest). The solution with the mushroom (Day #2) also turned darker over time, only faster.

Discussion:
a) The solution from Day #1 turned more yellow over time (the first cuvette was the lightest and the last cuvette, the darkest). This indicates that more product was produced over time. On Day #2, the mushroom turned darker faster, indicating that the reaction was occurring faster. This is probably because the mushrooms contain more enzymes, such as celliobiase, which can speed up the reaction to an extent. There is a limit to how many enzymes are effective, but in the case of Day #2, the extra enzymes did help speed up the reaction.
b) Possible sources of error...

• Incorrect measuring with the pipettes
• Contamination of the solution(s)
• Timing errors