Dicty transformation

http://www.nature.com/nprot/journal/v2/n6/full/nprot.2007.179.html

Electroporation
2 days prior assay, start shaking cultures: 0.3x10^6 cells/ml or previous day: 1.5x10^6cells/ml (~1 confluent plate)
Grow cells in shaking culture up to 4x10^6 cells/ml
Count 10^7 cells
Spin 1500rpm 3min
1x wash with cold Na/KPO4 buffer. (Resuspend pellet)
Incubate 15min on ice
Spin
Resuspend at 10^7 cells/ml in Electroporation buffer (1ml)
Mix gently 800ul cells with 30ug plasmid DNA (for linear DNA, start with 30ug plasmid, digest and clean)
Incubate 1min on ice
Transfer to cold electroporation cuvette and pulse twice (1kV, 3mF, 0.7-0.9ms) (within 5s)
Plate on 100mm dish with HL5 medium
Add selection next day: blastocidin 75ul/plate

Cloning on bacterial lawns
Resuspend cells in HL5, count cells
Resuspend ~2cm3 of plated bacteria (KA) in 5ml SM broth
Dispatch 250ul/ tube (2 for each transformant)
Mix 100cells in one and 300 cells in other
Spread on blue bar plates (SM)
Clones are visible in 3 days

Pick and inoculate in 1ml HL5.
Inoculate in 10mm plates when ready.