Weeks 6 and 7 were very similar. Kevin and I would observe the 6 tanks, each with 3 fish, for 3 hours. Then, we would remove the fish from the spawning tanks, weigh them, and put each fish back to their respective tank. Afterwards, Kevin went to the lab's office and enter our data for the day onto the computer. Simultaneously, I would remove the eggs from the spawning tanks, count the fertilized, unfertilized, and nonviable eggs in each tank, and set the eggs in the incubation tanks where they would grow for 24 hours.
The 24 hour mark is important to the experiment. After 24 hours, we are able to determine which cells were the result of the triploid or diploid male. This is because the DNA in the diploid zebrafish male's were altered to make the fish transparent. This specific type of zebrafish is called a "casper." As a result, the eggs laid by the casper zebrafish are also transparent. Due to this, we are able to determine the fertilization rate of each fish.
The fish to the left is a wild-type zebrafish, where as the fish to the right is an example of a casper zebrafish.
Kevin did a great job entering all of the data into the computer!
After gathering a great amount of data from my previous experiments and observations, the Professor decided to change the set up of the experiment. Instead of having two zebrafish in one tank (one male and one female), Dr. Dabrowski decided to follow the methods of another spawning experiment in the past; we began to have three zebrafish in each tank (one diploid male, one triploid male, and one female). We also decided to double the amount of tanks to increase the rate at which we get data. Since one person can't observe six tanks simultaneously, I received a new partner for the experiment: undergraduate student Kevin Fischer.
The protocol for observation was very similar to the previous weeks, so I was able to help introduce Kevin to the spawning observations we were looking for. Afterwards, we weighed and measured each fish. Then, Kevin and I would both check the fertility rate of each tank.
On August 27, instead of doing my usual 3 hours of zebrafish observations, I began my day helping with an in vitro fertilization. The Professor decided to discontinue the 3 hour observations because we had gathered enough information from them. Now, it was necessary to interpret out results and decide the next path to take.
To perform multiple in vitro fertilizations, it would be necessary to take the sperm from multiple different males; however, this is extremely inefficient. Instead, I took the testes from one male zebrafish, macerated the sample, and centrifuged the solution. Because of this, I had enough sperm for multiple fertilizations.
After I finished the in vitro fertilization, I helped feed the fish. The zebrafish's diet is brine shrimp (The orange solution).