Students will learn about Experimental Design by trying out a new, never-before-tested-in-yeast microscopy kit for Tetrahymena. The students will first make hypotheses and predictions for patterns etc based on the kit’s use in human cells and the differences with Tetrahymena and yeast. On Day 2, students will grow their cells, and then test several concentrations, settings on Day 3. Samples will be analyzed on Day 4, and data will be analyzed between Days 4 and 5 for a final report.
Students will clone a gene for use in the model organism Tetrahymena thermopile. Initially they will set up a PCR, and then clone into a vector for expression and grow up plasmid to clone into on Day 2. On Day 3, students will clone insert into plasmid through ligation and transmigration into E coli. On Day 4 students will screen colonies for correct ligation and then isolate several to grow overnight. On Day 5 they will mini prep the cloned plasmids and do a digest to screen for correct insertion.
University of Toronto Mississauga:
University of Toronto:
1. Mitophagy is a cellular process through which cells dispose of mitochondria that are dysfunctional or damaged. In this process, the mitochondria are progressively engulfed by a double membrane, thus forming an autophagosome. The autophagosome is then sent to lysosomes for degradation and recycling of its content. The student will use microscopy techniques to analyze the process of mitophagy in conditions of cellular stress.
2. TFEB is a transcription factor that normally resides in the cytosol. Upon induction of specific stress pathways, TFEB translocates to the nucleus where it interacts with DNA and induces the transcription of specific sets of genes implicated in the biogenesis of lysosomes that are important for allowing autophagy to take place. The student will use biochemical techniques (western blotting) and microscopy techniques to analyze the translocation of TFEB to the nucleus during infection with intracellular bacterial pathogens.
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