I have done competent cell preparation for cloning ( DH5 alpha) by Rubidium chloride method first time.But, by mistake the entire experiment I have accomplished outside the hood.Is it okay or I should discard those cells ?
I commonly make competent cells w/o use of a hood. However, when I do I always do the transfers/aliquots under the aid of a Bunsen burner/flame to sterilize the surrounding air. If no aseptic technique was used, I would discard and start again either in a hood, or use aseptic technique.
Ideally, you should prepare the competent cells under hood and using flame to sterilize the surroundings. This minimizes the chance of environmental bacteria contamination. You should take 1 or two trials with the competent bacteria you prepared, if you don't get any false colonies or opaque white colonies, you can retain them and vice- versa.
I also make competent cells w/o a hood, and I have never had a problem. Flame sterilization will help keep things sterile.
If you follow aseptic technique and your surrounding air are sterile then that is okay. Just give a trial of your prepared bacteria. if there is no false colony then there is no contamination. If you get any false colony then discard and prepared the competent under hood.
Best of luck
TA
The competent cells will be OK but try first transforming without and with known plasmid to make sure there is no contamination in the cells. We also make regularly competent cells without hood and in the presence of flame (to keep surrounding air sterile) and never had any problem of contamination.
- Badges
- Science topic
- Similar topics
- Biological Science
- Cell Biology