Electroporation of nec

From Tanaka Wiki
Revision as of 09:22, 25 March 2015 by Stephanh (talk | contribs)
(diff) ← Older revision | Latest revision (diff) | Newer revision → (diff)
Jump to navigation Jump to search


Culture media:

Name Quantity Remarks
Glutamax DMEM:F12
Glutamin 10µl/ml
Pen/Strep 10µl/ml
B27 (50x) 20µl/ml
FGF2 (20µg/ml) 20ng/ml
Shh (Curis) 100ng/ml For lower passages only


  1. Incubate NSC 60-90min with 1mM DPBS/EDTA
  2. Prepare 1.5ml tubes with DNA mix (max. 3µg/µl, 0.75-1µg/µl per plasmid). Max. 3µl DNA mixture
  3. Prepare 12well plate with gelatine
  4. Prepare Culture Media
  5. Centrifuge cells for 3min at 700g
  6. Resuspend one flask (75cm2) with cells after centrifugation in 160µl Steinberg solution
  7. Add 12µl each of the cell suspension to every DNA mix
  8. Electroporation conditions are: 500V, 50ms, 5 pulses
  9. Add cells to the plate and check after 1 day