Hey Everyone,
Today, unfortunatly the 10 different strain samples of propionibacterium acnes hadn’t grown enough to use yet, so we left them to grow longer all today and overnight.
We then removed the previously incubated old batch of plugs from SpeI enzyme, buffer and BSA. We incubated them all in 1 x wash buffer for 30mins at R.T. During that time we made up 1L of TAE 0.5x buffer, autoclaved it, allowed it to cool and used it to saturate the plugs for 1hr. We also used the buffer to make up a 1.4% gel, before cutting each of the 10 different strain propionibacterium acnes plugs from an old batch and placing them into wells. We also included a sample of the low range PFG marker. We then sealed the plugs using 1.4% agarose, we found a few problems whilst doing this. As we haven’t worked with as many plugs in the same gel at once before, one pipette tip (1ml) wasn’t enough to do all 11 wells. This was also made worst by how quickly the agarose cooled in the pipette creating bubbling and messy sealing. We plan to use a different tip for each sample next time, and work with slightly warmer agarose.
We then ran this gel for 18hrs, at 6V/cm, with 1-12 sec Switch interval. We also used the modified buffer flow method – which took a few times to balance effectively.
Tomorrow we will be hopefully starting plug production and viewing this latest gel. See you all then 🙂