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New to cloning -- strangely "mature" colonies after ON growth. Any idea


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#1 yoursisterdebra

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Posted 12 August 2010 - 08:32 AM

I am absolutely new to cloning so perhaps this is a dumb question... I did my transformation yesterday (E. coli competent cells) and plated on ampicillin plates around 4 pm yesterday. The plates sat at 37 degrees overnight and this morning at 9:30 am I checked them. My control plate was totally clean but my other plates had colonies on them that, according to our lab manager, were something she had not seen before. To her (she has been doing this for 12+ years), the colonies I got from overnight growth look more like what you would expect to see after 48 hours or more. They don't look like yeast or fungus, at least not grossly. I am doing further culture of them to see what happens but being inexperienced at this I am curious if anyone has an idea what I might be growing? Is it possible they are in fact my desired colonies but for some reason they grew at an increased rate? Thanks!

** EDITED at 6 pm **
Okay, I did a subculture in LB media + campicillin and when I checked after 6 hours they were visibly turbid. There was also substantial growth on the restreaked plates from this morning but it definitely doesn't look like E. coli. Still doesn't look like fungus or yeast either. Apparently I managed to culture a mystery ampicillin-resistant bacteria... joy.

Edited by yoursisterdebra, 12 August 2010 - 03:11 PM.


#2 perneseblue

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Posted 12 August 2010 - 08:37 PM

I am absolutely new to cloning so perhaps this is a dumb question... I did my transformation yesterday (E. coli competent cells) and plated on ampicillin plates around 4 pm yesterday. The plates sat at 37 degrees overnight and this morning at 9:30 am I checked them. My control plate was totally clean but my other plates had colonies on them that, according to our lab manager, were something she had not seen before. To her (she has been doing this for 12+ years), the colonies I got from overnight growth look more like what you would expect to see after 48 hours or more. They don't look like yeast or fungus, at least not grossly. I am doing further culture of them to see what happens but being inexperienced at this I am curious if anyone has an idea what I might be growing? Is it possible they are in fact my desired colonies but for some reason they grew at an increased rate? Thanks!

** EDITED at 6 pm **
Okay, I did a subculture in LB media + campicillin and when I checked after 6 hours they were visibly turbid. There was also substantial growth on the restreaked plates from this morning but it definitely doesn't look like E. coli. Still doesn't look like fungus or yeast either. Apparently I managed to culture a mystery ampicillin-resistant bacteria... joy.


Nice, if this bacteria grows very fast, maybe you might want to see if it can take up plasmids by transformation. It could be interesting. Transform in the morning, colonies in the afternoon and results by the evening.
May your PCR products be long, your protocols short and your boss on holiday




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