Electroporation of A1 cells
Jump to navigation
Jump to search
The printable version is no longer supported and may have rendering errors. Please update your browser bookmarks and please use the default browser print function instead.
Material
- Ice
- 4 mM cuvettes, N+1 cuvettes
- Steinberg's, at 4 C.
- Square pulse electroporator (we use the BTX 830 Squarporator)
Steinberg's
| 1x | FW (MW) | 5x/500 ml |
|---|---|---|
| 58 mM NaCl | 58.44 | 8.47 g |
| 0.67 mM KCl | 74.56 | 1.68 ml (1M) |
| 0.44 mM Ca(NO3) | 236.20 | 1.1 ml (1M) |
| 1.3 mM MgSO4 | 246.47 | 3.25 ml (1M) |
| 4.6 mM | Tris pH 7.8-8.0 | 5.75 ml (2M) |
Procedure
- Put cuvettes on ice, and cool down Steinberg's on ice
- Trypsinize cells as normal, stop reaction with high serum media and spin down at 1000 rpm, 3 minutes
- Wash pellet with Steinberg's solution and resuspend in 300 µl of Steinberg's, keep on ice
- Before electroporation, make sure to electroporate a cuvette with no cells to discharge device
- Electroporate cells in following conditions and put back on ice: Myoblasts: 100V, 35 msec, 5 pulses, Myotubes: 75 V, 35 msec, 5pulses
- Plate cells as normal