problem with autoclaving LB twice - need advice! (Jul/08/2009 )
I needed to make some LB-Amp plates today. Since I already have a lot of LB medium already, I decided to use some of that to make my plates.
I took 500 ml of already autoclaved LB broth (which I had made in November last year) and poured it into a 1 liter flask. I then added 7.5 grams of Agar-Agar, covered with Al foil and sent it off to autoclave.
When I got my flask back, I found that the LB-agar inside was a kind of darkish brown colour and also the volume was between 350-400 ml.
When i opened the al foil, I could smell a burning smell.
I added 400 ul of Amp and poured my plates anyway, but I am worried as to what the matter is.
Do you think my plates are usable or should I just discard them and make new ones again?
The media is burnt. Discard it and start over...
Come on lotus - you made the medium last November, reused in in making another medium and were apparently surprised that its appearance was off? Do you do any media controls - employ some concept of media expiration?
Amen to all of the above, but, another real problem was that you probably did not mix the agar well. It tends to form clumps and sit on the bottom of the bottle, then burn, since some of it is dry. Your autoclave may also have been set on a "dry" cycle, since you appear to have evaporated a fair amount of the solution.