Dear,
I make competent Agrobacterium with cacl2 and do transformation with them.
Finally I take colonies in plates but they are rough and dry and separate on lb agar and not mucilage?
Why it happen?
Hi, what Agro strain are you using. Did you check the presence of transgene in agrobacterium. I feel there may be some contamination. You can try to repeat the whole experiment with fresh chemicals.Also try to use fresh antibiotics.
We use Rifampicin to avoid contamination with other bacteria (usually E.coli); Agro is naturally resistant to Rif so you can use it together with the antibiotic selection for your plasmid.
Doesn't sound right. Probably you should do it again with another established protocol.
Could you post your protocol? Some Agro strains are more happy in YEB medium
Hello bolok, i have attached a file for the agro comp cell preparation by cacl2 method you can go throw that. you can use YEP or YEB media in place of LB.
Thanks a lot anybody
Mara Schuler : 3.2. attached file is this protocol i use it
Hi, there! As mentioned above, the genetic background of the strain is important. There are a lot of "tricks" available to improve efficiency for certain strains. If you give us your protocol we might be able tp "tweek" your procedure.
Best wishes,
Klaus
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