Direct Agrobacterium Transformation Of GV3101(PMP90RK) With PLH Vectors
Thaw the competent cells carefully on ice (here it took 50 minutes).
Add 0,1-1 µg of DNA to the cells and mix.
Freeze the cells at -80°C
Thaw the cells by incubating in a 37°C water bath for 5 min.
Place the cells on ice for 30 min.
Spread the cells on prewarmed LB plates (28°C) containing all the appropriate antibiotics
After 2 days incubation at 28°C about 20-100 colonies will appear.
http://www.dna-cloning.com/?q=agrobacterium
I think the amount of time it takes to thaw depends on the type of method used to prepare the competent cells. If the cell preparation does not contain 10% or more of glycerol or equivalent, then it will take longer to thaw. But that should not be a problem. I have worked with certain strains of bacillus that also used to take long to thaw.
Thank you very much for your comments. I got nice colonies after 2 day of incubation.
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