why do i need to use recA positive bugs for pGEX vectors?

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markbond
markbond's picture
why do i need to use recA positive bugs for pGEX vectors?

Recently tried cloning a gene into pGEX6P-1.  Cloning all went well but th vector did not express.  After a bit of head scratching I read that it is not recommended to clone and grow these vectors in recA negative bugs (e.g. DH5 alpha which I use).   I thought recA negative bugs were better for maintaining insert and vector stability.  Why do I need to use recA positive bugs for my cloning.  The tech support guy confirmed that this is the case but could not give a reason.

Anyone else come across this?

Thanks

Mark

varsha
varsha's picture
GE does mention that recA1

GE does mention that recA1 mutation containing strains DH5a and XL-1 are poor choices for GST fusion clones. In past I have used both DH5a and XL-1 blue for cloning of pGEX vector/insert. They worked fine for maintenance, for my inserts. However,  used BL21 cells for expression and purification of the fusion proteins.
You oculd take out the plasmid DNA containing your insert from DH5a and transform into BL21 for expression.  Good luck!
Varsha