Recommended competent cell for transformation of large plasmids? - (Mar/19/2012 )
Which competent cell type would you recommend for transformation of plasmid sized at 16 kb? My gene of interest is inserted into the pCrl empty vector.
From background reading, I have narrowed this down to JM109 and XL-10 Gold. Has anyone used either or both for large plasmids?
I've used neither- but we use either DH10B or DY380 (depending on application) for our BAC (which is about 230 kb).
DH10B should be okay ...watch out for strains harboring deoR and hte mutations (GeneHogs, XL10-Blue, Stbl4, SURE). I would use electroporation ...this should give you better transformation efficiencies for larger plasmids!
This was my question too. I'm about to transform an 18 k plasmid, but my TOP10 can't take more than 10 K. I always thought DH10B is the source of TOP10. don't remember where I heard this....so according to you guys DH10B should be fine, and I will try electroporation.
The 10Kb is not an absolute limit. The efficiency of transformation for longer plasmids is low, but you should be able to get a transformant. There are strains designed for accepting longer fragments. I think Lucigen and Epicentre are two companies that come to mind.