How to get rid of satellite colonies? - (Jun/10/2007 )
Ampicillin plates (100ug/mL) often show satellite colonies after incubating for more than 16 hours (my plates show satellites after 20 hours incubation). So it is better to incubate for a shorter period.
We had the same problem and switched to carbenicillin, while using the same vecotr, and no more satellites but colony growth.
I agree with the use of carb over amp and believe that the extra cost is worth it. Also, don't forget that amp is light sensitive and will quickly break down if you are storing your plates in even just a little bit of light.
What is your usual incubation time?
hi macedo,
somebody told me not to incubate plates(with amp.) too long. could you tell me why not to incubate plates no longer than 16 hours at 37℃？is it related the stability of amp.？
thanks.
What is your usual incubation time?
hi macedo,
somebody told me not to incubate plates(with amp.) too long. could you tell me why not to incubate plates no longer than 16 hours at 37℃？is it related the stability of amp.？
thanks.
in a sense. Beta lactamase is secreated into the medium. So for most e coli strain, after 16hr, there is enough beta lactamase secreated into the agar surrounding colonies which have grown up, to allow, satelite colonies (bacteria without the plasmid) to grow.
I would say that if you do get satellite colonies, this isn't too much trouble. Just streak out the central colony onto a fresh amp plate, It would give you your resistant colony by dinner time
I agree with the use of carb over amp and believe that the extra cost is worth it. Also, don't forget that amp is light sensitive and will quickly break down if you are storing your plates in even just a little bit of light.
Hi,
Is ampicillin really light sensitive? Because I was under the impression that solely the x-gal componenet of the plates was light sensitive.
Thanks!
Ampicillin is light sensitive but not as light sensitive as x-gal. It will deteriorate over time if you leave your plates in light. Always keep aliqouts rather than thawing large volume stock solutions on each usage as is every other stock that you have.
I had the problem of satelite colonies before as well where I then prepared fresh stock of Ampicillin for I was actually using a year old Ampicillin that was stored at -20 degress C.
Other than that, if you prepare your plates, don't leave them to solidify for long in your laminar airflow but rather take them out as soon as it hardens. Old plates are considered 3months in the chiller. So throw away plates that are old.