Examine whether the ubiquitination or sumoylation motives of Chmp1Aare functional in the binding and/or posttranslational modification with ubiquitin or sumo protein, respectively
My summer research is being conducted at the laboratory in the University Of Pikeville Kentucky College Of Osteopathic Medicine, KYCOM. When my research started I had no experience other than that from lab in chemistry and biology from school, but that comes nowhere in comparison to this, so my first week or two consisted of training from Dr. Maiyon Park. I learned the techniques of Cell culture using two cells lines Human Embryonic Kidney (HEK 293) and Pancreatic Cancer (PanC1, adenocarcinoma) cells. The techniques I learned with cells include splitting cells, transfection, protein isolation, cloning of Chmp1A mutant constructs using restriction enzyme digestion, gel purification, ligation followed by DNA purifications. For this project, I also use bright field, fluorescent, and confocal microscopic analysis, Western Blot analysis, in vitro binding assays and in vitro sumoylation and/or ubiquitination assays. My project is still going underway we are still collecting data, but my primary focus is the investigation of and proving of the existence of the Ubiquitin Interacting Motif and Sumoylation sites of Chmp1A, a known tumor suppressor. In order for us to study this we needed to make four constructs, which has turned into a quite lengthy process. A construct contains an insert of the DNA you want to investigate, UIM1 UIM2 SUMO Mutant 1, and SUMO Mutant 2 in our case. To make these constructs you have to use molecular cloning. I have my constructs made and I am in the process of finalizing my data.
Dietary Supplements as alternative therapeutics for pancreatic cancer patients
I have been doing collaboration with two medical students, so I’ve been tackling two research projects this summer. This project is to determine the benefit of dietary supplement as therapeutics for pancreatic patients. For this project, I learned to treat cells with dietary supplements, count cells using an inverted microscope and a hemocytometer, how to examine the changes in known tumor suppressor gene expression by Western Blot and confocal microscopic analysis, as well as spheroid formation assays, which is a 3-D growing environment. So far the data is looking promising, we have seen some noticeable changes in cell growth, death, and even increase in Chmp1A and pp53 expression which are known tumor suppressors according to our western blots and confocal microscopic analysis of the staining for these proteins. This data is still preliminary and needs to be repeated, which we are in the process of repeating our cell counting, spheroidal analysis and western blots, before we can state this as true but it looks like a good step in the forward direction of Pancreatic Cancer Research! I love doing this research and plan to continue doing research throughout my education.