Learning New Skills in the Lab

Sarah Ye is a junior majoring in Neuroscience. She was awarded a Spring 2017 Independent Grant which she used to conduct research on spermatogenesis and genome instability under Dr. Anthony Chan.

I have been working at the Yerkes National Primate Research Center in Dr. Chan’s lab for almost 2 years now. I started as a work study student my freshman year and I enjoyed going to Yerkes so much that I decided to take a research class for my second-year fall semester. Since starting at Yerkes, I have learned so much interesting information and learned many valuable skills. Right now, I am developing a transgenic mouse model for studying genome instability in spermatogenesis.

Since there is high chromosome modification when a spermatocyte turns into a spermatozoon the genome instability during spermatogenesis can be quite high. This genome instability may lead to mutations in coding genes which can result in diseases such as Kennedy’s disease, spinocerebellar ataxias, and Huntington’s disease. These diseases are caused by trinucleotide repeat (TNR) mutations in coding genes. Although germline expansions in trinucleotide disorders are well documented, the expansion mechanism is unknown. A working model system will be needed to further study TNR disorders and genome instability during spermatogenesis. My study aims to create a transgenic mouse model with a spectrum of fluorescent tags that are expressed in specific sperm cell types in the testicles. These “testi-bow” mice will allow sperm cells to be identified during a specific stage of development which can be isolated and sorted for molecular analysis to determine the genome instability of sperm cells at different stages of development. “Testi-bow” mice will be characterized based on their specific transgene expression pattern in the sperm cells, and will be central in subsequent studies investigating genome instability and TNR expansion.

Here I am looking at the embryos that I have injected with the lentivirus. It has been 4 days since I injected them and they have developed into blastocysts. 

Since starting my project I have learned that working with embryos can be very challenging at times and many steps need to be done correctly to collect the embryos. Before the embryos can even be collected the female mice need to be injected with hormones to stimulate ovulation. After hormone injection, they need to be mated with a male mouse. The timing is critical to this experiment because once mated the female must be checked for the vaginal plug in order to ensure successful mating. The fertilized embryos should be collected as soon as possible because the perivitelline injection (injection into the space between the cytoplasm and the zona pellucida) of the lentiviral vector should be done before the zygote undergoes its first division. During the days that I scheduled to collect embryos I needed to check the vaginal plugs and start collection around 7:00am otherwise the plugs would melt. Mice are nocturnal animals so assuming that the mice mated around 12:00am the plugs should be checked no later than 8:00am. It is also good to collect the embryos early to have enough time to do the perivitelline injection before the zygotes undergo their first division.

Here I am collecting the embryos form the oviducts of the mice. The embryos also need to be washed in an enzyme to remove the cumulus cells. 

Performing the perivitelline space injection is relatively straightforward but washing and transferring the embryo can be quite challenging. To perform the injection, the cumulus cells, which surround the embryo, need to be washed off. This is not easy since cumulus cells are very sticky and they are hard to come off. The embryos need to be put in the enzyme hyaluronidase for the cumulus cells to loosen. The embryos can’t be left in the hyaluronidase for too long otherwise the embryos will lyse. After detaching the cumulus cells the zygotes need to be transferred to microscope plate the do the injection. Transferring the embryos is also challenging because they are very small and it is easy to lose a few during the transfer. It takes a little practice to get used to transferring the embryos to different plates.

Even though embryo work takes time to get used to I am glad that I started this project. I have learned so many new skills and I hope to use them in order to execute and finish my project. I know that there may be many problems and challenges ahead but I will try my best to overcome them and reach my aim.

Visit the Undergraduate Research Programs website to learn more about applying for Independent Research Grants.