In 2015, we designed and synthesized 3 composite parts, but were only able to test one of them for proof of concept. This was because we did not succeed in cloning the other two composite parts into the yeast vector, thus we were not able to get the composite part into yeast and test it. This year, we decided to try again to get those composite parts into yeast. To do so, we changed the restriction enzymes and gel extracted the yeast plasmid (instead of a DNA purification). The gel extraction was to prevent the plasmid from ligating back on itself, without the insert inside the plasmid.
We succeeded in cloning one of our composite parts from last year into yeast, Composite Part (BBa_K1871002). We then tested the genetically modified yeast containing this composite part. We were able to confirm that the yeast were expressing the alpha amylase enzyme by showing starch degradation in both the short term (over a 6 hour period) and in the long term (over a period of 3 days). The genetically modified yeast containing this part was also tested in several samples of industrial water from a ceiling tile plant over several days, using a prototype to test the yeast. Lastly, this yeast strain was used in our cell growth testing, the results from which were simulated using the mathematical model we created. Our results are shown below. Figure 1. The image above was taken 5 hours after adding the genetically modified yeast to a starch solution. The yeast strain containing a plasmid with this composite part is the third tube from the left. The right most tube is the control, containing wildtype yeast and starch. Iodine, which reacts with starch to form a blue color, was added to each solution to determine the amount of starch. The third tube from the left has a very light blue color, indicating a small amount of starch left. The right most tube has a dark blue color, indicating a large amount of starch left. This shows that the yeast cells are producing and secreting amylase enzymes, thus the composite part is fully functional.Figure 2. The graph above shows a quantification of starch degradation over a period of 6 hours. A spectrophotometer was used to quantify the amount of blue color in each tube, seen in the figure above. An absorbance level of 1 equals a starch concentration of 0.25%. The yeast strain with this composite part is labeled as "Modified Yeast Strain 3" in the graph. For information on our results please see our Results page. With these new results, we were able to improve the characterization of this part. The newly characterized part can be seen on the Registry of Parts.
