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Host cells for transformation

transformation

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#1 Gunalan

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Posted 03 December 2011 - 11:37 PM

Hii all
Can any one suggest which is the best host cell for transforming pQE 30 vector with insert and pRSET A vector.
Also suggest me the source form which i can read more about these aspects please.


Thanks
Gunalan

#2 pDNA

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Posted 04 December 2011 - 07:10 AM

you can have a look at the Qiagen Expressionist Manual for the pQE30 ...for details see [url="http://www.qiagen.com/literature/render.aspx?id=128"]here![/url]

pQE30 contains the T5 promoter that is recognized by the E. coli host RNA polymerase (in contrast to the T7 promoter) ...generally you can use any E. coli strain for expression. I personally prefer the JM109 since it contains an F-plasmid with an extra lacI that gives you tight repression of the T5 promoter and prevents leaky expression. For a detailed list of available E. coli strains can be found [url="http://openwetware.org/wiki/E._coli_genotypes"]here[/url].

For the pRSET series of vectors you need a DE3 lysogen like BL21(DE3) ...since those plasmids use the T7 promoter.

Regards,
p

#3 Gunalan

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Posted 05 December 2011 - 12:56 AM

you can have a look at the Qiagen Expressionist Manual for the pQE30 ...for details see here!

pQE30 contains the T5 promoter that is recognized by the E. coli host RNA polymerase (in contrast to the T7 promoter) ...generally you can use any E. coli strain for expression. I personally prefer the JM109 since it contains an F-plasmid with an extra lacI that gives you tight repression of the T5 promoter and prevents leaky expression. For a detailed list of available E. coli strains can be found here.

For the pRSET series of vectors you need a DE3 lysogen like BL21(DE3) ...since those plasmids use the T7 promoter.

Regards,
p




Thanks a Lot pDNA for ur suggestions
It will be highly helpful to me
I am using E. Coli DH5 cells for bot vector transformation as on now I'am planing for a sub cloning later on
is this approach is correct
as now I don't have the other strains

#4 pDNA

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Posted 05 December 2011 - 03:28 AM

for vector transformation and plasmid preps DH5alpha is fine ...so you don't have to worry!

Regards,
p





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