Recently, I've been trying to transform Arabidopsis protoplasts with PEG method, but failed.
The plant is 3 week old Col in greenhouse, watered every other day.
The protoplasts look good under microscope, but I got no transformants with 35S::GFP at all.
I followed the protocol by Abel & Theologis from Arabidopsis Protocols (Humana Press, 1998).
My question is if there is any trick in the transformation procedure? Actually, I've tried
several protocols from different labs, but all the experiments failed.
Does the whole transformation process have to be operated under sterile condition?
Is there any factor considered critical or important for a successful transformation?
I really expect to learn from your advice if you have experience with such transformation.
Thank you very much in advance for your helpful reply!