Is it ok to replate from a bacterial lawn?

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Is it ok to replate from a bacterial lawn?

Post by Newbie » Fri Jun 09, 2006 3:01 pm

Hi all,

I was doing some plating on LB plates and let it grow overnight. The next day I noticed that I had a lawn of bacterial growth, probably becuase I plated the entire transformation mix and it was way too concentrated. Anyhow, I was just wondering whether it would be ok to replate from this bacterial lawn, and just streak some onto another plate or whether I should re-do the transformation and plate less volume this time. Also, if I do re-plate from the bacterial lawn how much of it should I plate? I was thinking of transferring some colonies into 1ml of LB and then taking 100ul of this to replate. Would that be too much still?

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canalon
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Post by canalon » Fri Jun 09, 2006 5:33 pm

Plating a full transformation? You were a bit pessimistic about your experiment, weren't you? I usually plate 100µl of the 1ml SOC, and it is usally enough ;)

Anyway, I would say that it would probably be better to restart from the beginning. But it depends on your selection: if ampicillin, definitely restart from scratch (it gets degraded and you will probably have a majority of non plasmid carrying strains). If you have a stable antibiotic, like kanamycine, you can try to just scrape some bacteria and replate them on selective medium. Of course it works if you have only one insertion, not if you are trying to have a library...
Patrick

Science has proof without any certainty. Creationists have certainty without
any proof. (Ashley Montague)

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Post by Newbie » Mon Jun 12, 2006 6:11 pm

Hi canalon,

Yea, I was a bit afraid of my low concentration of DNA. I will re-do the transformation then! Thanks.

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