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Curcuma longa and Wild Type p53 Gene Induced Apoptosis in MDA-MB-231 Breast Cancer Cell Line Objective and Application to Biotechnology ` Conclusion Results and Interpretation Eight million people die every year due to cancer. This research will be a step forward in treating cancer, saving those lives. This experiment will focus on treating cancer using antioxidant, specifically Curcuma longa, and p53 gene. This comparative investigation will draw parallels and contrast the effectiveness of antioxidant and p53 gene individually and both treatments combined. Such treatment can be easily implemented since it will not harm normal cells but will selectively kill cancer cells. If an antioxidant is added to the cancer cells, then oxidative stress will be reduced. This will activate the inserted p53 gene. Hence, cell death will be initiated by the antioxidant and inserted p53. Figure 2: Free radicals are damaging the cell. Figure 1: Free radicals are produced through several day-to-day activities. Materials and Methods Figure 3: Images of cells with plasmid DNA before and after the experiment under a fluorescent microscope. Figure 4: High levels of oxidative stress deactivate the p53 protein and activate NF-kB protein which makes the cell cancerous. ` or s. f t ial a e tr p e Re or m ➢ Obtain p53 Plasmid ➢ Count Dead Cells ➢ Grow Single Colony ` ➢ Obtain DNA ➢ Culture Cells Figure 5: The graph shows the average percentage of cell death by all the experimental groups and the controls. The data shown is an average of three trials. The error bars show the respective standard deviation of each variable. Untreated cells had 0% cell death; hence, no error bar is shown. With that, p-values by T-tests are shown. * signifies p <= 0.05, ** signifies p <= 0.01, and *** signifies p <= 0.005. ➢ Transfect DNA Abstract and Personal Information Variables & Controls Dependent Variable: Dead Cells (%) Experimental Groups Constant: ` ● ● ● ● ● ● Amount of PBS Amount of Trypsin Amount of media Amount of Trypan blue Amount of DNA Etc. ` Controls Eight million people die every year due to cancer. Hence, this experiment will focus on treating cancer using Curcuma longa and p53 gene. This comparative investigation will draw parallels and contrast the effectiveness of the antioxidant and p53 gene individually and both treatments combined. Furthermore, a wild type p53 plasmid was bought from Addgene. Then, a single colony was isolated after overnight growth. This colony was grown for another 24 hours in LB Broth with Ampicillin antibiotic. Then, miniprep kit from Qiagen was used to isolate plasmid DNA from the bacteria. It was transfected into MDA-MB-231 breast cancer cells using Lipofectamine 3000. Then, Curcuma longa (an antioxidant) was added after a day. After three days of incubation, cells were counted using Trypan blue and a cell counter. In conclusion, Curcuma longa activated the inserted p53 protein and may have deactivated the NF-kB protein. Thus, apoptosis was induced in MDA-MB-231 breast cancer cell line. Curcuma longa with p53 protein killed 56.67% cells on average. On the other hand, Curcuma longa and p53 gene individually killed only 53.00% and 6.33% of cells respectively. The T-test confirms that most experimental groups were significantly different from the controls. Vraj Shroff, Niles West High School, Skokie, Illinois Antioxidant initiates cell death in cancer cells by altering transcription of proteins that induce apoptosis and prevent cells from proliferating forever. Cancer cells are very different from the normal cells because of the high level of oxidative stress. This inhibits the activities of p53 gene, hence apoptosis is avoided. With that, NF-kB protein is overly expressed, which allows cancer cells to proliferate infinitely. However, antioxidants reduce oxidative stress in the cells and reverse this situation. Curcuma longa activates the inserted p53 gene and deactivates the NF-kB protein. Thus, apoptosis is induced in MDA-MB-231 breast cancer cell line by Curcuma longa and p53 gene. The results confirm the hypothesis of this investigation. Curcuma longa and p53 protein killed 56.67% cells on average. On the other hand, Curcuma longa and p53 gene individually killed only 53.00% and 6.33% of cells respectively. The T-test confirms that two experimental groups are significantly different from their controls. The p-value of p53 DNA with Curcuma longa to Control DNA with DMSO is 0.002. While p-value for Curcuma longa to DMSO is 0.023. Lastly, the p-value for p53 DNA to Control DNA is 0.321. Hence, two of them are well under 5% similarity. The data also suggests that experimental groups may be different from each other as well. The p-value for Curcuma longa to p53 DNA is 0.004, and the p-value for p53 DNA with Curcuma longa to p53 DNA is 0.005. However, the p-value for p53 DNA with Curcuma longa to Curcuma longa is 0.461, meaning more trials are needed to find a statistical difference between those two experimental groups. Acknowledgements ` I am very grateful to Niles West High School for providing me an opportunity to conduct my independent experiment. I want thank District 219 Township for providing lab equipment and grants to carry out this research. I acknowledge the efforts of science faculty, namely: Ms. Swiercz, Mr. Ruber, Ms. Gleicher, Mr. Kim, and Ms. Villa. With all the teachers, I want to thank Ms. Arcus for providing all the materials required for this experiment. Lastly, I am indebted to Ms. Schwarz, my mentor, for providing help around the clock. This experiment may not have been possible without her.