21 Mysteries

Hello all!

Here goes my final blog post for the summer…

In order to select for reporter activity, I identified colonies that were growing on the quadruple selective plates (- TRP-URA-HIS-ADE). After performing the yeast two-hybrid screen, I had a total of 21 blue colonies on all of the plates! Each blue colony represents a protein that interacts with E6-AP. I have been home in California and away from lab, so I have not yet extracted these proteins and then sequenced them in or der to find out what they are. I cannot wait to head back to lab in a few days to discover what these proteins are! Hopefully I will find new protein interactions with E6-AP.

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2 Hybrid Screen

Hello All!

I finally performed my yeast two-hyrbid screen a few days ago!

I crossed my bait, the protein E6-AP, with my prey, the human fetal brain cDNA library in it. I had to prepare a concentrated overnight culture of my bait strain. The next day, the library strain was combined with the bait strain. Then the cells were put in a shaking incubator over night. After 24 hours, I plated my mated culture. Plating the mated culture was an ordeal, because I had to pipette 100 μl of the mated culture on each plate. Mind you, I had plate fifty 150 mm plates. These plates were missing certain amino acids, in order to select for true interactions. The yeast will take 5-8 days to grow. In my last blog post, I will let you know how many colonies grew!

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The Real Excitement Commences

Hello all!

Last time I blogged, I had successfully made my clones to work with! So as a review, I have my gene of interest, inserted into the plasmid that contains the binding domain and that construct is now in a specific yeast strain. I then looked through our lab’s database to see what genes people have cloned into the activation domain. After finding around 20 genes that others have cloned into the activation domain, I performed a separate transformation for each of the 20 genes, into the yeast strain containing my gene of interest, UBE3A (Also called the bait).  I then plated the transformants onto selective media and put the plates into a 30 degree incubator, where they will grow for 3-5 days. After the 3-5 day growth period is up, I will take a single colony from each plate and streak it onto another selective plate. If colonies grow after 3-5 days on the new selective media that indicates that a TRUE PROTEIN INTERACTION is occurring! I made a diagram that shows what I talked about, unfortunately, I could not get it to go directly on my blog post, but if you go into my gallery and click on “Pictorial Explanation of 3rd blog,” you can see the diagram! Here is the link- http://ccsummerresearch.blogs.wm.edu/files/2012/08/In-Yeast-Strain1.pdf

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Fun with cloning!

Hello All!!

 

So, I arrived in Williamsburg on June 3rd and promptly started lab research on June 4th. Time really does fly when you are fully engrossed in a research project.  Just as a re-cap on my project. I am performing a Yeast Two-Hybrid Assay, to find proteins that interact with the protein, UBE3A. I could go into every nitty gritty detail of each step of the process, but I feel as if I would completely bore you. So, I am going to explain it in a simplified way. A scientific experiment in a wet lab takes many many many steps. Each step takes much longer than what would be expected, especially when working with yeast.

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Research on Angelman Syndrome

Hello all!

My name is Sophie Cohen and I am honored to receive one of the Chappell Fellowships. I am 6 days away from finishing my sophomore year at the College!! After all of my exams, I am going to go back home to Scottsdale, Arizona for a few weeks, before coming back to Williamsburg to conduct research. I am a Neuroscience Major and am going to try to complete a Hispanic Studies Minor.

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