I have been having some new issues with seeding NGM plates. The lawn looks fine, but when you look under the microscope you see these tiny dots/colonies growing on the lawn. When left out another day overnight they end up growing and becoming these larger viscous white spots but only stay on the e.coli lawn, they don’t grow anywhere else on the agar plate. I added strep and made sure to decontaminate the space, so if it is some sort of contamination I’m assuming it’s coming from the plate that has the single colonies I pick to inoculate.
Does anyone have an idea what this could be? Could it be that I left the plates out for too long/gave the bacteria too much time to grow on the plate?
I am starting over from the glycerol stock and new LB media, so I’m hopeful it should be gone with the next plates I seed. But I’m just more curious if others have seen something similar to this.
If it’s only within the lawn, it seems likely it’s a contaminant within the culture you’re using to seed the lawn. If you really are picking single colonies to seed your plates, then it would have to be present in the culture medium. You could try leaving your culture medium at 37 for a while and seeing if it becomes turbid, and using this 37C-incubated media that you never inoculated with OP50 to spot some plates. You could also try streaking out from one of your mottled lawns, to see if more than one sort of colony arises.
The fun alternative would be that you’re only seeing this effect on the lawn because it’s a rare effect happening to the bacteria, something throughout the plate that causes a phenotypic change at very low frequency. I could imagine incredibly slight contamination with phage might be able to do it, though I haven’t heard of that.
I wonder if you happened to identify the cause for this ? My lab facing the similar situation. We’re working with OP50, so no antibiotic is use. After seeded on NGM, and leave at room temp for few days, we started observe these dots and only grown on the lawn.
Do you have photos of the spots? I had people tell me revertants may cause that. OP50 is engineered to not grow too well, and sometime revertants may occur and form spotted colonies. My lab solved it by getting a new vial from CGC