Thesis is fully underway!
The goal of my project is to understand the role of Nfe2 and how it affects the development of neuromast cells (supporting cells) in the inner ear under oxidative stress.
The basic idea of my project is what effect does diquat have on inner ear development when NFE2 is expressed normally, and not fully expressed.
Last week in the lab was very productive yet challenging. It started with running qPCR for three of my primer I will be using. Initially, the results were not what I was hoping for but as Carolyn Lawson always use to tell me “data is data”. We attributed this issue possible to a lack of cDNA in my dilution series. However, with the great help of Anna Marie, I was finally able to produce qPCR that had appropriate Ct and standard deviation values to move on in this process! I also got some more practice at microinjection zebrafish embryos. For my particular project, I will be injecting Brn3c embryos, which have green fluorescent protein (GFP), which enables us to image their nueromast using confocal microscopy. This week I injected with control morpholino, which has no effect on the expression of NFE2.
This week has been a relatively busy week in the lab. Lab this week started with me and Anna Marie arriving at the lab at 5 a.m. on Monday to dose our zebrafish embryos. Why so early in the morning you ask? At this time, our Brn3c embryos that have been injected with control morpholino are 92hpf, in which has been designated as the time in development to dose the fish. We then come back at 9:00 a.m. to image the 96hpf embryos in order to determine the distance between otoliths, length, and width of the inner ear. Later this week I will be running qPCR to determine the housekeeping gene for my qPCR project! Currently, I have chosen 18s and b2m as potential candidates for my housekeeping gene. I will also microinject embryos this week, continuing to use my control morpholino!