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BL-21 Gold (DE3) cells and further strains


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#1 samita

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Posted 10 February 2011 - 09:09 AM

I was preparing competent cells for few strains but it was with out antibiotics. Now I am using BL21-Gold (DE3 and), Codon Plus-Chloramphenicol, and PLYS strains--Chloramphenicol. When Codon plus and Plys strains are used for expression, I used amp and Chlor both together. Which antibiotic is used when prepare competent cells of all these strains?????. Tetracyline or Chloramphenicol???
regards

#2 pDNA

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Posted 10 February 2011 - 09:53 AM

during preperation of competent cells you want to asure that both plasmids are present in your cells ...therefore i would add both antibiotics present on the plasmid backbone ...in your case, as far as i get it, Chloramphenicol (CAT) and Ampicillin (AMP).
You wrote CAM and TET (tetracycline)? ...why?

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p

#3 samita

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Posted 10 February 2011 - 11:15 AM

Amplicilin is used for your expression vector like pET32 for expression of protein...which is not there when you are preparing the competent cells.
But BL21 Gold (DE3) is modified version of BL21 cells and resistant to tetracycline. I am little confused now in competence cells preparation and during expression of protein. When cells are resistant to Tetracycline, I can use it inorder to avoid from contemination. But do I need to used Tetracycline during expression of protien from vector which have Amplicilin gene as well.




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