I'm now using Gene Pulser Xcell (Biorad) to transform a vector (pUC118) into E.coli DH5α. In fact, I did this procedure many times and got satisfying rssults. However, I encountered a problem these days. I usually used the pre-set protocol for E.coli with parameters as follows, 1.8kv, 1mm, but after pulse, the actual output was only 14v. I think that's the reason why I can't get any recombinats on the LB plate, but I don't know the cause and the way to solve it. Can someone help with that? Thanks a lot.
If you are only getting 14V output then that most certainly is the problem. The problem most likely is with the machine, but it could also be with the cuvettes or with the competent cells (such as the what solution they are in). First make sure everything is plugged in well and the connections are good (especially the cuvette holder) and that it is clean so the electrodes are making good contact with the cuvettes.
You can then try some control experiments, just add ddH20 to your cuvette instead of cells and see what sort of voltage you get. If it is still not correct, borrow a new cuvette (preferably a different brand) from some other lab and try that. If that still is not working, then there likely is a problem with the Gene Pulser itself.
If those work well but your actual electroporation is not, then what solution are using to do the final cell resuspension? And when you apply power is there a loud noise that is made (which happens when there is too much salt)?
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