I have a two-pronged plan to get a phage strain that gives good enough plaques for my GTA-as-vaccine experiments.
I obtained reasonable titers of two phages, 'Titan' and 'Saxon'. I'll invest a couple of weeks to see if I can get better and more reproducible plaques with either of these. The genome sequences of these phages are not closely related.
First, improve the plaquing conditions: The researcher who isolated the phages recommends using for the lawn cells that have been grown photosynthetically to a high density, He also suggested trying a lower top-agar concentration. I'll play around with these and other variables to see if I can get better plaques.
Second, use artificial selection to get a better strain of phage: I'll pick the few best-looking plaques of each of my two phages and plate the phage they contain in new lawns. From those new lawns I'll again pick the best-looking plaques, and plate their phage in new lawns. Etc. Maybe I'll introduce a bit of UV mutagenesis along the way.
The first step will be to make fresh lysates of these phages. The lawns I made before are too old, so I'll grow up some cells for lawns today and tomorrow I'll retiter the lysates. On Friday I can pick plaques from these lawns and make plate lysates. If there's a plate with near-confluent plaques I can use it directly to make a plate lysate. (10^7 or 10^8 pfu/ml, and I have maybe 5 µl so at best I can get plates with 5 x 10^4 or 5 x 10^5 plaques. The latter might be enough to get a good lysate. There are small volumes (50-100 µl?) of the original lysates in the lab upstairs, so maybe I'll sue these. Or Maybe I should save these until I've improved the plaquing conditions.
RFK Jr. is not a serious person. Don't take him seriously.
3 weeks ago in Genomics, Medicine, and Pseudoscience
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