I am doing bacterial plasmid transformation on the newly purchased plasmid example FoxP3 merge with 10B competent cells. We used LB agar with ampicillin and inoculate the said mixture through it. After following our protocol without missing parts of it.On the second day, we noticed there is the growth on the LB agar and we proceed to stage two by making grow in LB broth with ampicillin on 10 ml tubes but after 6 hours, we found out no turbidity on the said broth but other tubes shows positivity due to presence of turbidity so we decided to throw it away, we never repeat what we did on day 1. I am new in this field, My question is what cause why there is no growth shows from tube we inoculated while other has? Is there something wrong with the protocol or the product we buy from the manufacturer. Anyone with great expertise with this field, tell me what happens on here. I need your help. Thank you
Maybe your plasmid don´t have resist to ampiciline, or your protocol to transformation is incorrect. You can tried this protocol: https://www.ncbi.nlm.nih.gov/pubmed/6345791
First of all I couldn't understand which broth showed growth and which didn't, please explain clearly.
Secondly, the colonies on agar plates may not be the transformant colonies so, they didn't grew in LB broth. There might be some problem during transformation process.
Next, you need to check your said plasmid and competent cell details before proceeding further.
Good luck!
I agree with the comments given by Divya Das. Your question does not look clear. You need to post clear questions to get the right answers and appropriate comments and suggestions to fix your problems. What are your controls? How did you perform your transformation? What methods did you use for your transformation? Heat shock? electroporation? others?
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