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e.coli contamination


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#1 deespike

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Posted 04 June 2010 - 09:14 AM

Hi all

I am having some contamination problem with my e.coli competent cells, i am getting white colonies after transformation.But i think dh5alpha cells donot look white but translucent could that be fungus contamination? When i am checking them for plasmid i get nothing sometimes and sometimes there is rna only but no plasmid.

Has anyone has had same problem?
plz help.

thanks in advance

#2 pDNA

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Posted 07 June 2010 - 11:38 AM

hard to interpret from a far

do these colonies form within overnight incubation at 37C? ...or do they need longer?

are these self-made competent cells or do you bought them?

I would suggest its a prokaryotic contamination ...but as mentioned above ...hard to say without more information!

Regards,
p

Hi all

I am having some contamination problem with my e.coli competent cells, i am getting white colonies after transformation.But i think dh5alpha cells donot look white but translucent could that be fungus contamination? When i am checking them for plasmid i get nothing sometimes and sometimes there is rna only but no plasmid.

Has anyone has had same problem?
plz help.

thanks in advance



#3 zowawih

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Posted 21 June 2010 - 11:21 PM

I had a similar problem.
I repeated the insert ligation and the transformation as simple as that.
Some times its hard to visualize the plasmid after the extraction coz the plasmid can spread to different light bands represent different forms (Supercoiled, liner ...etc). To turn around this, you have to find out your plasmid size, the size of the insert and calculate these numbers together, then digest the plasmid by restriction enzyme and run it on an agarose gel. You should visualize a band with the same size of your plasmid + insert.




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