I am going to use CRISPR/Cas9 system for knockout a gene. CRISPR is in the form of a plasmid for an idividual gene. My question is that can I propagate CRISPR plasmid by transforming E.coli ? If anyone is have experienced the same situation please help me in this.
Yes you can propagate it as long your plasmid has prokaryotic origin of replication.
Thank you Ashim but that information is not available I only know that it has U6 promoter.
U6 promoter is for expression of gene in mammalian cells since finally CRISPR will go to human cell line. But please focus on my question whether it is possible to propagate in bacteria for future use and endless production in own lab.
As Aasim Majeed answered already, if the vector have prokaryotic origin it can transforming in E.Coli and propagate. Usually vector have the Ori and selection marker to propagate. if you want know more information, we need to know vector is from where.
Hi Anuj, I have propagated CRISPR Plasmid ( px330 ) in Stbl3 as well as Dh5 alpha comp cells, so the answer is yes.
Dear Anuj Kumar Singh
Since this discussion was done several years ago,
however, I want to know that whether you became succeeded to propagate Crisper cas9 plasmid in competent E. coli, that was targeted to eukaryotic cells?
I am stocked at the same point where you year at 5 years back, If you did it, please let me know the detail about it.
thank you
- Badges
- Science topic