pGLO Observations , Data Recording & Analysis

pGLO Observations , Data Recording & Analysis

1.

Obtain your team plates.  Observe your set of  “+pGLO” plates under room light and with UV light.  Record numbers of colonies and color of colonies. Fill in the table below. Plate Number of Colonies Color of colonies under room light Color of colonies under   UV light - pGLO LB carpet gray gray - pGLO LB/amp 0 gray gray + pGLO LB/amp 104 gray gray + pGLO LB/amp/ara 1/2 carpet/ 64 gray green (glowing)

2.	What two new traits do your transformed bacteria have?
	My transformed bacteria glow in the dark and are resistant to ampicillin.
3.	Estimate how many bacteria were in the 100 uL of bacteria that you spread on each plate. Explain your logic.
	I think about 90 bacteria, because E. coli are about 1-2 micro liters each, and there were 100 micro liters.
4.	What is the role of arabinose in the plates?
	The arabinose made the bacteria glow by inhibiting the promoter and allowing the DNA polymerase to read the GFP Gene.
5.	List and briefly explain three current uses for GFP (green fluorescent protein) in research or applied science.
	Three current uses for GFP are to study cancer in mice, to remove malaria gene in mosquitoes, and to study the spread of HIV in cells.
6.	Give an example of another application of genetic engineering.

Crops that are resistant to herbicide.

Candy Electrophoresis Lab Analysis

Candy Electrophoresis Lab Analysis

1. When we analyzed the results of our gel, we found that none of our sample dyes contained different dyes than in the four reference dyes.  However, the dye band from our blue M &Ms was bigger than the Blue 1 reference dye. Also our green Mike & Ike band was much smaller than the yellow 6 reference dye. Our brown M&M had a multicolor band, with both red and blue. None of our dyes moved in the wrong direction. 

2. The Blue 1 reference dye would migrate similarly to the carminic acid, because they have similar structural characteristics.  The Red 40 reference dye would migrate similarly to the Betanin, because they both are bonded in similar patterns. The Blue 1 reference dye would migrate similarly to the Fast green FCF, because they are very very similar in their structure. The Red 40 reference dye would migrate similarly to the citrus red 2 because their structural characteristics are alike in many ways. 

 3. Dog food Manufacturers might put artificial colors in dog food to make the food look more appealing to the dog owners. 

5. The two factors which control the distance the colored dye solutions migrate are charge and size.

6. The electrical current helps move the dyes through the gel. 

7.  The small holes in the gel allow the smaller DNA strands to move quicker through tot he end with the positive charge. The bigger strands have a harder time going through. 

8. I expect the molecule which weighs 600 daltons to go the farthest, and then 1000, then 2000 (probably covering half the distance of the 1000) and then the 5000 very very far behind. 

Recombinant DNA Lab Analysis

Recombinant DNA Lab Analysis

In this lab, we made "recombinant DNA" models out of paper. Recombinant DNA is created through a process called transformation. First, a "gene of interest" is located, finding the location and sequence of the gene and its surrounding sequences. In our case, this gene was called the insulin gene. During transformation,  Enzymes called Restriction Enzymes cut the DNA when it reaches a specific sequence of bases. Different enzymes cut at different sequences. We used an enzyme called Eco RI, because it cut our plasmid once and our DNA twice, close to the gene of interest. This is important because it allows us to splice in the insulin gene to the plasmid. If we used an enzyme which cut the plasmid in two places, then the plasmid would be split into two pieces and we wouldn't be able to put in the gene. When it cuts it, it leaves a "sticky end," which helps the DNA bond with other DNA or plasmids. Plasmids are rings of DNA most commonly found in prokaryotes. Usually they have genes in them which give antibiotic resistance. Our plasmid contained a gene for resistance to both Tetracycline (used to treat acne and skin infections) and Kanamycin (treats serious bacterial infections). Ligase, an enzyme which puts base pairs back together, bonds the cut plasmid and DNA. Then the recombinant plasmid and bacteria are mixed, and the non-resistant bacteria are weeded out by adding whichever antibiotic the recombinant plasmids are resistant to.  For example, we would add in either Tetracycline or Kanamycin, because if the bacteria have taken in our plasmid, which has resistance to both, than it will survive. If it doesn't, it won't. We wouldn't use an enzyme such as ampicillin, because our plasmids are not resistant to it.  After the mixing, the gene is extracted and purified. This process is important in our everyday life because it allows us to mass produce necessary items such as insulin and more. This process could be used for herbicide or pesticide resistant crops or to clone organisms as well.

This picture is of our "recombinant plasmid" and the different restriction enzymes.

New Year Goals: 2016

New Year Goals: 2016

My first goal for the semester is to have neater, more thorough work in Biology.  I often rushed on assignments in first semester, and had terrible penmanship throughout. This worked against me in the long run, when I went back to study my completed notes and labs, I couldn't read them. I will achieve this by taking more time on my notes and being more careful when writing my lab reflections and relate and reviews.

My second goal for the semester is to to become better at balancing my schoolwork and extracurricular activities. To do this, I will focus more on my work my working in my room at my desk, and I will bring my smaller work with me in the car. I will make a schedule and plan my time according to practices and free time.

Unit 5 Reflection

Unit 5 Reflection

In this unit we learned about the Dogma. First we learned about DNA, which is made up of nucleotides. Nucleotides have three parts- the nitrogen base, phosphate group, and sugar.  DNA is anti parallel, and codes for every trait that you have. To replicate, DNA "unzips" and DNA polymerase rematches it- thus creating two new identical strands, consisting of the bases A,T,C, and G. We also learned about protein synthesis, which consists of two parts- transcription and translation. During transcription, DNA "unzips" and RNA polymerase matches all the spare nucleotides to make an RNA strand- messenger RNA. The RNA code replaces the T with a U. After it is produced, it leaves the nucleus and goes to the ribosome.  Once there, the ribosome reads the messenger RNA in groups of three bases (called codons) and translates it into the language of proteins (amino acids). Third, we learned about mutations, and how they effect proteins. The two main types of mutations are frame shift mutations (insertion and deletion) and mutations involving substitution. Finally, we learned about Gene expression. We learned that every cell in your entire body had the exact same DNA, but not all are expressed. There are different parts which control gene experession, including the promoter, the operon, and the operator. 

I'm good at translating the DNA to RNA and the RNA to amino acids, but I have a little difficulty understanding gene expression and the different parts. I am a better student today because I can now translate DNA to RNA to Amino Acids very very easily and quickly. I also learned better studying techniques, including making tests for myself and hearing the questions out loud instead of reading them. I also made flashcards on certain topics for practice. This works a lot better for me because I am a hands on or listening learner.  

I would like to learn more about how DNA and RNA polymerase work and what other molecules are involved in the unzipping and re matching of DNA. I have no unanswered questions. I wonder about how scientists first discovered protein synthesis and DNA replication, as it happens on such a small scale.

https://en.wikipedia.org/wiki/DNA_polymerase

DNA polymerase and the unzipping of DNA

https://en.wikipedia.org/wiki/Amino_acid

Finished Amino Acid Chain

Protein Synthesis Lab

Protein Synthesis Lab

There are there two parts in the creation of a protein: transcription and translation. During transcription, DNA "unzips," and RNA polymerase re-matches those spare nucleotides, thus making an new RNA strand: messenger RNA or mRNA for short. The messenger RNA leaves the nucleus and travels to the cytoplasm. There, it finds the ribosome. The ribosome reads and translates the RNA 3 bases at a time (these are called codons). It translates it to protein language, or amino acids. Each codon is one amino acid. These chains of amino acids are folded up to make proteins.

nyu.edu/classes/keefer/story/story5.htm

Based on what I have seen, frameshift mutations - both the insertion or deletion of a base - seemed to have the greatest effect on the proteins, as both completely alter the sequence of the bases. When we inserted in a C base to the DNA strand, it mutated the protein so greatly that there was no end and most of the amino acids were changed. When we deleted a T base from the DNA strand, it dramatically shortened the chain of amino acids. The location of the mutation does matter, because if we had deleted a base near the end of the chain then the outcome wouldn't have been as dramatic. The same goes for insertion; if we had inserted a base near the end, then the resulting protein wouldn't have been effected as substantially. The substitution of a base seemed to have the least effect on the protein. When we substituted a C for a T in the DNA strand, there was no effect, as the codon which was changed still translated to the same amino acid. The location of the substitution does not matter, unless the mutation occurs in either the first or last codon- thus changing the start or stop amino acid.  

I chose deletion because in my previous experiment it had the greatest effect on the protein. This mutation didn't alter the protein quite as severely, but still resulted in a protein with no ending. It does matter where the mutation occurs, and if a base closer to the beginning of the amino acid chain had been deleted, then the resulting protein would be almost completely different than the intended version. 

Crohn's disease is a disease caused by frameshift mutation- specifically the insertion of a C base. It is an inflammatory bowel disease. This disease causes you to lose a lot of weight, have pain in your gut,  have a lot of dietary restrictions, and much more. These are symptoms of the disease because it causes swelling in the digestive track as well as ulcers (holes).  

Human DNA Extraction Lab

Human DNA Extraction Lab

In this experiment we questioned whether DNA could be separated from cheek cells in order to study it.  We found that it could be separated, as long as the procedure is followed extremely carefully. It can be observed after precipitation. We know this because we extracted our own DNA, directly after adding the alcohol to our DNA solution. The DNA was white and floated at the top of the test tube.  We knew it could be observed after precipitation based on our prior research.  This data supports our claim because after adding the alcohol we immediately saw the effects on the gatorade solution.

While our hypothesis's supported by our data, there could have been some errors due to timing mis calculations and due to step mix ups. My mixing up the stepping we could have prevented precipitation and messed up the whole experiment. By mis calculating the time we could have also messed up the solution and caused the alcohol to have no effect. Due to these errors, in future experiments I would recommend using a timer instead of watching the clock, and triple checking your procedure.

This lab was done to demonstrate how DNA can in fact be removed by your body cells, and how it is present on every cell that you have.  From this lab I learned the process on DNA extraction which helps me understand the meaning of precipitation.   Based on my experience from this lab, I could apply this extraction technique to other uses such as on fruits like strawberries and bananas.