Gene Researcher at the Taubert Lab: Day 4 & 5

I spent my last two days at the Taubert lab extracting plasmids from bacteria. I followed the alkaline lysis procedure in which bacteria is exposed to NAOH and sodium dodecyl sulfate (SDS), a strong detergent. The cell membranes of the bacteria burst and the substances inside the bacteria spill out. Sodium acetate, an acidic solution, is then added to neutralize the solution. I observed that by this point, the cell membrane material and DNA material have precipitated. I then separate the cell contents by centrifugation. The resulting substance is extracted to purify the plasmid DNA.

I used a spectrophotometry too. It is a photometer that can measure intensity as a function of the light source wavelength. It is used commonly used for the measurement of transmittance or reflectance of solutions, but it can also be used to measure the equilibrium constant of a solution.

I have learned so much this week at the lab. I can’t wait to work in a genetics lab in the future!!

- Alice Yip

Gene Researcher at the Taubert Lab: Day 3

Today, I learned about Western blot also known as the immuno blot. Just as PCR detects and amplify specific DNA sequences, the western blot detects specific proteins in a sample. I once again used gel electrophoresis to separate proteins by 3-D structure or denatured proteins by the length of the polypeptide. This time I used SDS- Page as in most proteins, the binding of SDS to the polypeptide chain gives an even distribution of charge per unit. I also heated the sample to 60 degrees Celsius to promote protein denaturation and thus, help SDS bind. A tracking dye was added to the protein solution so we could track the progress of the protein solution through the gel during the run. Afterwards, the proteins are transferred to a membrane where they are searched using antibodies specific to the target protein. Since it was my second time making gel electrophoresis, I found easy to pipette the solution into the holes.

 

Although reading about Western blotting does not take that long, the actual process actually took the whole day. And in the end, only one of the two samples worked. I was a bit disappointed, but I guess that is what happens in science: experiments don’t always turn out the way you want them to turn out.

- Alice Yip

Gene Researcher at the Taubert Lab: Day 2

Today, I experimented with gel electrophoresis, a method used to separate proteins by charge and size. In this experiment, it was used to separate DNA fragments and proteins. Nucleic acid molecules are broken apart by applying an electric field which transports the negatively charged molecules through an agarose matrix. As I was observing the electrical field, I noticed that shorter molecules move faster than longer ones and I concluded it was because shorter molecules travel more easily through the pores of the gel. For this procedure, I followed the QIA Quick gel extraction and the buffer I used was TAE.

 After the electrophoresis is complete, I stained the molecules in the gel to make them visible. I was able to see them under ultraviolet light while using protective gear, of course. I took a photo, but the color of the DNA turned out to be an unattractive green and the gel was black. I could not even recognize the green parts were DNA when I took the photo. I guess this sheds light on another side of genetics and it illustrates to me how DNA does not look “beautiful” in real life.

While waiting for the gel to thicken, I made bacterial and seed cultures and synchronized worm growth in C. Elegans.

Another day at the lab has passed by without me even realizing it until now. I wonder what I will be doing tomorrow!

- Alice Yip