1. Equipment: Gene Pulser (Bio-Rad).

2. Phosphate-buffered saline (PBS): 150 mM NaCl, 2.7 mM KCl, 4.3 mM Na2HPO4, 1.4 mM KH2PO4 (pH 7.4); sterilize through a 0.2-^m filter and store at 4°C.

Fig. 1. Electroporation efficiency of adipocytes tested using pcDNA3.1-LacZ. Differentiated 3T3L1 adipocytes were electroporated with 0 or 600 ^g of pcDNA3.1-LacZ, and transfection/expression efficiency was determined by X-Gal staining for b-galactosidase expression.

3. Complete medium: Dulbecco's modified Eagle's medium (DMEM) plus 10% fetal bovine serum (FBS).

4. Dulbecco's PBS without Mg2* or Ca2+ (D-PBS) (Gibco-BRL, cat. no. 14190-029).

5. 15- and 50-mL conical tubes.

9. Cuvet stand.

10. Collagen-4 coated six-well plates (Fisher Scientific, prewarmed at 37°C).

11. Plasmid DNA (CsCl doubled banded).

12. Trypsin-EDTA (0.05% or try 0.25% solution). Warm to room temperature prior to use.

0 0

Post a comment