Discipline: Biological Sciences
Subcategory: Cancer Research
Demia Jones - Harris-Stowe State University
The question that I am researching is whether or not there are circadian rhythms in gene expression in tumors. More specifically, if there are circadian rhythms in gene expression in Bmal1 knockout tumors. To test if these Bmal1 knockout tumors have daily rhythm in vivo, I measured mRNA levels taken from tumors at 4 different times of day (6am, 12pm, 6pm, and 12am). I began by isolating and purifying mRNA from brain tissue and tumors. After extracting mRNA, I measured the 260/280nm and the 260/230nm optical density ratios to ensure that the mRNA is not contaminated. Then I performed cDNA synthesis, which synthesizes DNA from the RNA template, via reverse transcription using a Polymerase Chain Reaction (PCR). The combination of reverse transcription with PCR will amplify the amount of sample available for analysis. I had to determine the best annealing temperatures for my Per2 primers using the cDNA that was generated from the brains of the mice, so I tested the primers on a standard curve as well as a melting curve to determine the efficiency of them. I then used qPCR to quantify the amount of Period2 (Per2) mRNA from each sample at the four times of day when the tumors were extracted. We hypothesized that in vivo, Bmal1 knockout tumors, express daily rhythms in Per2 mRNA levels compared to the housekeeping gene, GAPDH. We found that Per2 is in fact circadian in vivo, and we concluded that daily rhythms in the host (e.g. hormone levels or body temperature) drive daily rhythms in the Bmal1-null tumor cells. We noticed that the expression in Per2 nearly doubled during the day. This suggests that we can treat these tumors according to the time of day of the host.
Funder Acknowledgement(s): National Science Foundation (NSF); National Institute of Health (NIH).
Faculty Advisor: Erik Herzog and Tommie Turner,