crestwind24 – This morning I am screening through many, many plates of worms. Usually this is very tedious, but today it is very exciting because I am looking for worms that have had their genomes edited using the CRISPR/Cas9 system!!! The CRISPR system allows scientists to make specific genetic changes in the actual genes of bacteria, cells, and animals. In the case of C. elegans, the CRISPR system allows scientists to target genes within the genome to mutate them, change them, delete them, or add things to them. It is extremely powerful, and hopefully I will find some worms that have the genetic edit that I designed!! Cross your fingers for me!
psgurel– Speaking of tedious, I am in the final steps of expressing and purifying protein. Purified protein is a requirement for a variety of biochemical experiments (like the kinetic assay or analytical ultracentrifugation I mentioned earlier). This whole process takes 5 days for the protein I’m expressing, where I’ve had to: grow and express my protein in E. Coli (E. Coli is typically used for protein expression because they grow very fast and yield a lot of protein), lyse the bacterial cells, harvest the protein through various methods of chromatography, concentrate my protein, then store it appropriately. Here I am posing with the FPLC, in the final step of purifying my protein. Looks like I got a good yield, yippee!
psgurel: I always save my favorite experiments for Fridays! Today, I’m doing an assay to look at kinetics. ATP (Adenosine triphosphate) is typically known as the main “energy” source in cells and is required for several reactions to take place. On a chemical level in these reactions, ATP gets hydrolyzed and you are left with ADP (Adenosine diphosphate) and phosphate as a product. Today, I’m looking at how fast different proteins hydrolyze ATP. To do this, I stop reactions at various time points, and then I add a green dye that labels free phosphate. The dye turns darker shades of green as the amount of phosphate in solution increases. Check out my samples! The darker the green, the more phosphate is present, which means more ATP has been hydrolyzed!
crestwind24: Today I am scoring and imaging the neurons of old worms. This means sitting for a few hours in a dark room at a big microscope and computer. Specifically, I am trying to see what is happening with certain neurons in the worm as they grow old. We know very little about how neurons change in old age, and why some die in normal aging, as well as in many diseases. In the first picture you can see the green light under the microscope, which is activating my fluorescent proteins. On the computer screen you can see the image the microscope is taking. The arrows point to the 2 worms I was taking picture of, old worm selfie!! The selfies are a bit grainy since the room is dark… sorry … and its time to upgrade my phone.
crestwind24: Today I am doing PCR to amplify a gene I am interested in from the DNA of a worm (C. elegans). PCR, or Polymerase Chain Reaction, can be used to make DNA, check for the presence of DNA, and/or sequence DNA. PCR is commonly used to detect the presence of viruses, like Ebola, by looking for the DNA of the virus in the patient’s blood or bodily fluids. Setting up a PCR involves pipetting small volumes of liquid, containing DNA, enzymes, salts, etc into tiny tubes (see images). Then the tubes are placed into a cycling machine (ours is named Cycle Jackson… get it?) that changes temperature over and over in a cycle to activate the enzyme that makes the DNA. At the end I hopefully will have a bunch of the DNA I want!
psgurel: Today I am staining grids for Negative StainElectron Microscopy. I will be using the TEM (Transmission Electron Microscope) which essentially uses a high voltage electron beam to visualize samples. My samples are placed on tiny grids (the arrow in my picture) and then stained with uranyl acetate, which is slightly radioactive (hence why I have to wear a lab coat) and scatters the electron beam. As a result, my samples stained with uranyl acetate will not absorb electrons and thus I can visualize them in contrast to the grid surface which will absorb electrons. Why use TEM instead of other types of microscopy? A typical fluorescence microscope yields about 200nm resolution. However, I’m trying to visualize protein clusters of less than 100nm in length…about 10,000x smaller than a grain of sand! The TEM will be able to resolve these structures!