Hi everyone! This week I finished my aPKC GFP probe and tested it for the first time on Thursday! It has been incredible to work through all the steps necessary to create a probe and then to end up with something that could be tested. One part of being a scientist that I am discovering is that sometimes what you are testing works perfectly with your hypothesis; but other times you can end up with a result that doesn't. While the probe did express on Thursday it didn’t begin to show up visibly until the eight-cell stage and even then, the probe was hazy. We were hoping to see it clearly between cells. The two places that the probe did show up clearly were in the nucleus and in small circles that we think could be vesicles. Unfortunately, neither of those were places where we wanted to see it! So, the next step we tried was to increase the amount of RNA that we inject into the embryos. On Friday we injected three times as much RNA as we did the day before. The embryos seemed to handle the higher dosage just fine, but the same dimness and lack of defined location in the cell was still observed and the probe was still not well defined by the time that cell polarization or first cleavage happened. However, when we looked at the cells that we injected on Thursday the probe was working. We could see it in the cell junctions. So, the probe is working, it just isn’t working at the stage in cell development that I need it to. There is a possibility that the probe only shows up briefly during first cleavage, so we just aren’t catching it when it is lighting up. I will be continuing to take videos of the injected embryos to see if we can see anything happening.
This week I thought that I would show you what I am looking for when I am watching the barnacles. Image 1 is a barnacle searching for a mate. The long thin projection is the penis that he will probe the other barnacles with as he tries to find a barnacle close to him to fertilize. Image 2 is a successful fertilization. You can actually see the thin thread of sperm as it moves from the donating barnacle to the receiving barnacle. Image 3 shows another way to tell if a barnacle has been fertilized. It will release a cloud of excess sperm 10-20 minutes after fertilization. The pink is from the colored pencil that we use to keep track of the barnacles. After I see a mating or a release of sperm I wait until the animal’s feeding appendage is out again before retrieving the brood. Then I pull apart the brood so I can inject the embryos.
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AuthorHi! My name is Sadie and I just graduated from Central Oregon Community College in Bend Oregon. I am working in Dr. von Dassow’s lab and I am excited to learn about research and cells. Archives
August 2019
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