Hello,
I am doing some bacterial transformations, and I need a GFP plasmid for reconstruction. I need to insert a new promotor region into the plasmid, and then heat shock it into an e. coli strain.
Does anyone have a recommendation for a good GFP plasmid vector? I am still designing the primers for the isolation of the promotor region, so restriction enzymes can vary (preferably EcoR1 though).
Thanks,
Hi Adam,
This plasmid was working very well as a positive control in our experiments:
http://parts.igem.org/Part:BBa_I13522
There is an EcoRI site upstream of the promoter sequence, but I would recommend replacing the promoter via Gibson assembly. If you are still working on the primers, you could think about it. It might be easier than finding a second restriction enzyme for a classical cloning approach.
Good luck!
Hello, thanks for your suggestion. If I was to use regular DNA recombination, do you know of any kits that could make the digestion and ligation procedure easier?
Thanks.
Hi Adam, You may also use Plasmid: pcDNA3.1/CT-GFP Fusion. It is easy to transfect in the bacterial system with CMV promoter.
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