Virginia Tech/infect

The next scale of our model involves matching infection of bacteria by phage λ

This was simple because we had generated plenty of growth curves of bacteria and had produced numerous phage lysates. All that needed to be done was run the experiment with both bacteria and phage, then interpret the data.

We wanted to test the OD595 of LE392 in the plate reader when the amount of LE392 and λ-EYFP is varied. Thus, we began by growing an ON culture of LE392 and generating conversion from OD600 from the spectrophotometer to the OD595 of the Plate Reader.

We then decided to run the different dilutions of LE392 and the different MOIs on one plate. The plate had dilutions of LE392 at OD595s of 0.029, 0.0435, 0.058, and 0.0725. The plates also had MOIs of 0.25, 0.5, 1.0, 2.5, 5.0, 10, and 25 of λ phage. The plate was placed in the reader to read the OD595 every five minutes, while shaking and maintaining a temperature of 37oC, for a total of 125 time intervals. MilliQ water was added after the first 25 cycles to prevent dehydration and then the plate reader was allowed to run for another 100 cycles overnight.

Results

Interpretation

The results are still being interpreted at this time. It is interesting to see the overall data produced by this experiment. High MOIs instantly killed the majority of the cells, but lysogens began growing quickly. Lower MOIs eventually killed the cells, but towards the end it was possible to see that some lysogens may begin growing. There was some obvious errors during the experiment. At one point during the time in the plate reader a blank well got inoculated accidentally. There might have been some human error as well; a couple of the wells appeared not to have been infected when they should have been. Next steps

We are going to run a similar experiment in black plates to get fluorescence data and run a parallel experiment in a clear plate to get absorbance data.