- inoculate 10 ml lb + kan50 + rif30 + gent15 with gv3101 agrabacterium containing your vector. grow agro culture at 28-30ºC.
- once culture is saturated (usually 24 hrs with fresh agro; 2-3 days from freezer stock), subculture into 800 ml lb + antibiotics. grow o/n.
- harvest cells by centrifugation; 5’ at 5000k.
- resuspend cells 800 ml of 5% sucrose.
- once resuspended, add 200 µl silwet, mix well, and transfer cells to a clean tupperware container.
- dip plants into agro solution. lay pots on their sides in a tray. cover with plastic wrap overnight.
- remove plastic and set pots upright. put them in the plant room away from other transgenic plants.