DAY 10: 19/06/17
The western blot prepared on DAY 9 was ran and scanned. Image is shown below:
The results didn't show a clear protein purification and that could be, for instance, while loading the gel, solutions ran from one well to another, mixing things up. However, it was unclear why the protein wasn't fully purified. Independently of the results not being clear, it was a good practice to make a western blot myself as I had never done it before (at uni, when we used one we were given a 'bought' ready one).
Today, spores solutions for b3.4 and 3 transformed plasmids were prepared and incubated overnight at 32oC
DAY 11: 20/06/17
Today, the spores solutions prepared yesterday, DAY 10, were plated out on EMM + Histidine + Adenine plates, and all 16 solutions (4 colonies from each plasmid) from plasmids 1, 5, 8 and 10 were plated on YE4S plates (YE4S medium had ampicillin and chloramphenicol added to it, in order to avoid bacterial contamination).
INFO: YE4S medium is a rich medium, and therefore all fungi should grown in it. EMM, however, as mentioned before, is a minimal medium and only the cells adapted to overcome the nutrients deprivation in the medium will survive.
So, for now any further step in my experiments require waiting for results, waiting for cells to grow. Thus, tomorrow will be a day off. Whoop, whoop I will sleep much much later than I'm used to these past weeks! 😎
DAY 12: 22/06/17
Today all plates were analysed to see colony growth. Also, because I've been using so many plates, I've prepared some more YE4S plates, and also YE4S + 5FOA plates for future use.
Again, in order to proceed with my experiments, waiting time to allow cells to grow more was required, so the weekend will be pro-longed with tomorrow! 😁
DAY 20: 24/07/17 The results from DAY 19's PCR is shown in figure 16. It can be seen from figu...
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DAY 5: 12/06/17 Today's plan was to confirm whether plasmids 3 and 7, in fact, contained the insert at the correct orientation. This wi...