Introduction: The microarray has become a new research tool for technicians looking to view and interpret genes. It is essentially a merge of genomics, computer science, and nanotechnology. It allows for a detection of patterns or changes in transcription in normal and abnormal cells. For example, you can compare cancerous and normal cells easily. Basically, a DNA microarray chip is a solid matrix, like a glass slide, that is imprinted with arranged pattern of spots that each represent part of a genome.
Procedure: 1)Prepare the simulated microarray slide
*put drop appropriate gene solution onto each slide
2) Hybridized your microarray with labeled cDNAs from normal lung tissue and lung cancer tissue
*label normal with blue dye, cancerous with red dye
*drop 20 uL of hybridization solution onto each spot
3)Visualize your labeled mircroarray results
Results:
Thursday, December 23, 2010
Wednesday, October 27, 2010
CSI: Lafayette. The Crime Lab
A) Viruses are able to inject their DNA into another organism to infect it. However, many bacteria have developed restriction enzymes to work as a defense mechanism. Restriction enzymes cut specific base pairs of DNA. We can use this process to our advantage as a way to observe DNA. Through agarose gel electrophoresis, where the DNA segments are put into gel which is placed in a buffer solution. When we apply a current through the solution, the DNA separates. This is because the DNA has an overall negative charge, and thus is attracted to the negative side of the slab. This will separate the DNA segments by size because the smaller ones travel further.
B) The ability to isolate and indentify DNA has incredible utility in the real world. For example, in crime scenes, using human remains we can indentify whose they are. We can also study the differences between healthy and cancerous patients by comparing their DNA. Or study the DNA of a species or population.
Procedure: There are five suspects to a crime that has been committed. We must find out who committed the crime. We must mix samples of DNA with the restriction enzyme mix. The incubate the samples in 37 degrees Celsius water. Then centrifuge the samples in an agarose gel in the electrophoresis apparatus. Then place the samples in the gel and run the electrophoresis for 30 minutes.
Results: Chloe committed the crime.
B) The ability to isolate and indentify DNA has incredible utility in the real world. For example, in crime scenes, using human remains we can indentify whose they are. We can also study the differences between healthy and cancerous patients by comparing their DNA. Or study the DNA of a species or population.
Procedure: There are five suspects to a crime that has been committed. We must find out who committed the crime. We must mix samples of DNA with the restriction enzyme mix. The incubate the samples in 37 degrees Celsius water. Then centrifuge the samples in an agarose gel in the electrophoresis apparatus. Then place the samples in the gel and run the electrophoresis for 30 minutes.
Results: Chloe committed the crime.
Sunday, October 17, 2010
Wednesday, October 13, 2010
The Search for Biofuels
Introduction:
A) Biofuels are quickly becoming an important alternative to fossil fuels. The basic requirement for a biofuels is that it comes from biomass. The first generation of biofuels were produced from food products such as corn and wheat. However, this was criticized because to took food away from the human population. Thus the second generation biofuels came from byproducts of food production. It converted much of the waste into fuels. The third generation of biofuels comes from algae. To break down biomass to produce ethanol, you must use enzymes. Enzymes speed up the rate of chemical reactions and are generally proteins. The reactant in an enzyme catalyzed reaction is caled the substrate. Enzymes speed up the rate of a reaction by reducing the activation energy needed for a reaction. Cellulose is found in the cell walls of plants, and is broken down by cellulases into glucose. By using cellulases, the biofuel industry can convert the cellulose into more easily usable forms, such as glucose. They then take the sugar and convert it into ethanol by microbial fermentation.
B) The purpose of this lab is to explore how cellulose from cell walls is broken down into glucose by enzymes. It has relevance to the research industry because they want to find an easy way to break down biomass into more usable forms of energy, such as glucose.
C) In this lab, we will be testing the effects of enzymes on a reaction. We will fill five test tubes with an artificial substrate (p-nitrolphenyl glucopyranoside)) and then add our enzymes (cellobiase). At specific timer periods we will add a base to stop the reaction and to act as an indicator to see how much product we got. In day 2 we will repeat this except we will use mushroom extract as our enzymes.
D) The control is this lab is a test tube with substrate but without an enzyme. The variables are how long the reaction occurs before we stop it with a base. I predict that the longer the reactions occur, the more product there will be.
Procedure: http://www.youtube.com/watch?v=v8TeTsnlKrg
Discussion: The data we received made sense. Looking at the five test tubes, they went from a light yellow (for the shortest time) to a more dark yellow (for the longest time). As the reaction went for a longer time, more product was produced. When we used mushroom extract, we had a similar effect. It was hard to distinguish if the mushroom extract was a faster enzyme that the cellobiase we used on the first day. Possible sources of error in this lab could be that if the reactant ran out during the time, then the reaction would stop. It did not seem to run out because the later test tubes each got darker. However, if we continued timing, at a certain point, the reactant would run out.
A) Biofuels are quickly becoming an important alternative to fossil fuels. The basic requirement for a biofuels is that it comes from biomass. The first generation of biofuels were produced from food products such as corn and wheat. However, this was criticized because to took food away from the human population. Thus the second generation biofuels came from byproducts of food production. It converted much of the waste into fuels. The third generation of biofuels comes from algae. To break down biomass to produce ethanol, you must use enzymes. Enzymes speed up the rate of chemical reactions and are generally proteins. The reactant in an enzyme catalyzed reaction is caled the substrate. Enzymes speed up the rate of a reaction by reducing the activation energy needed for a reaction. Cellulose is found in the cell walls of plants, and is broken down by cellulases into glucose. By using cellulases, the biofuel industry can convert the cellulose into more easily usable forms, such as glucose. They then take the sugar and convert it into ethanol by microbial fermentation.
B) The purpose of this lab is to explore how cellulose from cell walls is broken down into glucose by enzymes. It has relevance to the research industry because they want to find an easy way to break down biomass into more usable forms of energy, such as glucose.
C) In this lab, we will be testing the effects of enzymes on a reaction. We will fill five test tubes with an artificial substrate (p-nitrolphenyl glucopyranoside)) and then add our enzymes (cellobiase). At specific timer periods we will add a base to stop the reaction and to act as an indicator to see how much product we got. In day 2 we will repeat this except we will use mushroom extract as our enzymes.
D) The control is this lab is a test tube with substrate but without an enzyme. The variables are how long the reaction occurs before we stop it with a base. I predict that the longer the reactions occur, the more product there will be.
Procedure: http://www.youtube.com/watch?v=v8TeTsnlKrg
Discussion: The data we received made sense. Looking at the five test tubes, they went from a light yellow (for the shortest time) to a more dark yellow (for the longest time). As the reaction went for a longer time, more product was produced. When we used mushroom extract, we had a similar effect. It was hard to distinguish if the mushroom extract was a faster enzyme that the cellobiase we used on the first day. Possible sources of error in this lab could be that if the reactant ran out during the time, then the reaction would stop. It did not seem to run out because the later test tubes each got darker. However, if we continued timing, at a certain point, the reactant would run out.
Wednesday, September 22, 2010
DNA collection and extraction lab
Introduction: Deoxyribonucleic acid or DNA is the building block for all life. Every living thing has DNA to store genetic information. It is responsible for determining many traits in a person, like hair, skin, and eye color. It also gives cells directions to perform all of the functions that are needed in an organism. The DNA structure is known as a double helix, because of the two spiraling strands that run anti parallel to each other. The two stands are held together by bases; the four bases are A (adenine), G (guanine, T (thymine), and C (cytosine). The bases are connected to a sugar and a phosphate group to form a nucleotide. Also, the base A always pairs with T, and G always pairs with C. These four bases are organized into your genetic material, and make messages for cells that are called genes. These genes code for the construction of proteins, which serve as the basic functioning unit for the body. In humans, our DNA sequence is 99.9% similar, and the .1% causes each individual to be unique. DNA is found within the nucleus of a cell. DNA wraps around proteins to form structures called chromosomes. Humans have 23 pairs of chromosomes, and the chromosomes make up all your genetic information in your genome. While DNA holds the directions for making proteins, the genes do not actually directly make proteins. Cells use ribonucleic acid or RNA to transfer parts of the DNA sequence to the ribosomes to make proteins.
Purpose: The purpose in this lab is to precipitate DNA.
Procedure: 1) Label tube with full name
2) Gently chew cheeks in order to loosen cheek cells
3) Rinse mouth with saline solution. The saline solution serves to neutralize the negatively charged DNA.
4) Spit solution back into tube
5) Add 2 ml of lysis buffer to break open and dissolve cell membrane; it dissolves the cell membrane by dissolving the phospholipids. The lysis buffer acts in a similar manner as dish soap and detergent.
6) Gently invert tube 5 times
7) Add 100 ml of protease, which is an enzyme that breaks down proteins. The specific protein we are looking to destroy is DNase, which breaks down DNA.
8) Gently invert tube 5 times
9) Place tube in water bath at 50 *C for 10 minutes to speed up the reaction and help break open the cell membrane.
10) Pour 10 ml of cold alcohol into tube, this action starts to precipitate the DNA.
11) Let tube sit for 5 minutes
12) Invert tube 5 times
13) Transfer DNA into necklace
Results and Observations: The DNA was formed inside the tube. The DNA never precipitated until I added the cold alcohol.
Discussion: The lab worked very well for me. I got a good amount of DNA to precipitate. Adding the lysis buffer and protease allowed me to get access to the DNA, and then protect the DNA from DNase. Then, the cold alcohol served to precipitate the DNA out of solution. Some possible sources of error in this lab could have been not getting enough cheek cells. If you shake the DNA solution to hard, it could cause the DNA to go back into solution.
Purpose: The purpose in this lab is to precipitate DNA.
Procedure: 1) Label tube with full name
2) Gently chew cheeks in order to loosen cheek cells
3) Rinse mouth with saline solution. The saline solution serves to neutralize the negatively charged DNA.
4) Spit solution back into tube
5) Add 2 ml of lysis buffer to break open and dissolve cell membrane; it dissolves the cell membrane by dissolving the phospholipids. The lysis buffer acts in a similar manner as dish soap and detergent.
6) Gently invert tube 5 times
7) Add 100 ml of protease, which is an enzyme that breaks down proteins. The specific protein we are looking to destroy is DNase, which breaks down DNA.
8) Gently invert tube 5 times
9) Place tube in water bath at 50 *C for 10 minutes to speed up the reaction and help break open the cell membrane.
10) Pour 10 ml of cold alcohol into tube, this action starts to precipitate the DNA.
11) Let tube sit for 5 minutes
12) Invert tube 5 times
13) Transfer DNA into necklace
Results and Observations: The DNA was formed inside the tube. The DNA never precipitated until I added the cold alcohol.
Discussion: The lab worked very well for me. I got a good amount of DNA to precipitate. Adding the lysis buffer and protease allowed me to get access to the DNA, and then protect the DNA from DNase. Then, the cold alcohol served to precipitate the DNA out of solution. Some possible sources of error in this lab could have been not getting enough cheek cells. If you shake the DNA solution to hard, it could cause the DNA to go back into solution.
Tuesday, August 31, 2010
Introduction
Yogurt is a popular snack that is created by bacteria. Bacteria are very successful, microscopic life forms. They are so small that they cannot be seen with the naked eye, yet they make up more of the total biomass than any lifeforms. There is a great range of types of bacteria, and many can be helpful for humans, and some can be very dangerous. Some examples of benficial bacteria are probiotics, which are found inside the human body and can help digestion and even lower cholesterol. We use many kinds of bacteria to create food and drink because bacteria go through a process called bacterial metabolism or fermentation. During bacterial metabolism, bacteria break down sugars into other chemicals through enzymes. From this process, we make wine, beer, and yogurt. However, harmful bacteria such as typhus, tuberculosis and the bubonic plague have killed millions of people throughout history. All bacteria share some common traits, for example, all bacteria are prokaryotes. The three major shapes that bacteria take are coccus (spherical), bacillus (rod-shaped), and spirillum (spiral).
In this lab, we will make yogurt, practice microbial technique, and test koch's postulates.
Procedure: 1) Label four tubes of milk as
Tube 1 - negative control (only milk)
Tube 2 - positive control (milk and yogurt)
Tube 3 - Yogurt and amp
Tube 4 - E. coli
2)Add 10 ul of ampicillin to tube "Yogurt and amp"
3) Dip fresh inoculation loop into the yogurt and swirl loop into "positive control"
4)Dip same loop into the yogurt again and swirl into "Yogurt and amp". Then put loop into bleach beaker
Observation: Tubes "negative control", "yogurt and amp", and "E. coli" all stayed as milk. Tube "positive control" had yogurt qualities (ie: smell and to some extent texture) but it was not fully formed as yogurt.
Discussion: My hypotheses were correct. The only tube that formed yogurt was "positive control", because it had yogurt bacteria and a hospitable place to cultivate. "Yogurt with amp" did not turn into yogurt because the ampicillin killed all the bacteria in the tube. Neither tube "negative control" or "E. coli" had any yogurt bacteria, so they did not turn into yogurt. One possible source of error in this lab is cross contamination. If some ampicillin got into another tube, or if yogurt bacteria got into a tube where it wasn't supposed to be, the results would have been skewed. Another source of error is not putting enough yogurt bacteria. Our "positive control" didn't fully form yogurt possibly because we did not have enough yogurt bacteria to begin with.
In this lab, we will make yogurt, practice microbial technique, and test koch's postulates.
Procedure: 1) Label four tubes of milk as
Tube 1 - negative control (only milk)
Tube 2 - positive control (milk and yogurt)
Tube 3 - Yogurt and amp
Tube 4 - E. coli
2)Add 10 ul of ampicillin to tube "Yogurt and amp"
3) Dip fresh inoculation loop into the yogurt and swirl loop into "positive control"
4)Dip same loop into the yogurt again and swirl into "Yogurt and amp". Then put loop into bleach beaker
5) Using a fresh inoculation loop, transfer E. coli to the tube "E. coli"
6)Place tubes in a an incubator for 24-48 hours
Tubes one and two act as control tubes, while tubes three and four are the variables. In this experiment, I believe that tube one will not change, tube two will turn into yogurt, tube three will not change (due to the bacteria killing ampicillin), and tube four will not change
Observation: Tubes "negative control", "yogurt and amp", and "E. coli" all stayed as milk. Tube "positive control" had yogurt qualities (ie: smell and to some extent texture) but it was not fully formed as yogurt.
Discussion: My hypotheses were correct. The only tube that formed yogurt was "positive control", because it had yogurt bacteria and a hospitable place to cultivate. "Yogurt with amp" did not turn into yogurt because the ampicillin killed all the bacteria in the tube. Neither tube "negative control" or "E. coli" had any yogurt bacteria, so they did not turn into yogurt. One possible source of error in this lab is cross contamination. If some ampicillin got into another tube, or if yogurt bacteria got into a tube where it wasn't supposed to be, the results would have been skewed. Another source of error is not putting enough yogurt bacteria. Our "positive control" didn't fully form yogurt possibly because we did not have enough yogurt bacteria to begin with.
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