Technical Reference #1902

1902.

Use of a microscope stage-mounted Nickel-63 microirradiator for real-time observation of the DNA double-strand break response Zhen Cao; Wendy W. Kuhne; Jennifer Steeb; Mark A. Merkley; Yunfeng Zhou; Jiri Janata; and William S. Dynan, Medical College of Georgia, Nucleic Acids Research, Advance Access(1902), (2010)
Link To Paper

Materials & Methods:
Cells and electroporation conditions- U2OS 2-6-3 cells (8) were maintained in high glucose DMEM supplemented with GlutaMAX-1 (Invitrogen) 10% fetal bovine serum 100 U/ml penicillin 100 mg/ml streptomycin and 0.25 mg/ml Amphotericin B. Electroporation was performed using 3 mg of plasmid DNA and 40 mg of sheared salmon sperm DNA carrier (Ambion Austin TX USA) using a Gene Pulser II (BioRad Hercules CA USA). Electroporation was performed in a 0.4-cm cuvette containing 120 ml of culture medium with settings of 170V and 975 mF resulting in a 60–90 ms time constant. Following electroporation cells were seeded into glass bottom culture dishes (MatTek Ashland MA USA) and incubated at 37 C to allow protein expression. Irradiation and imaging were performed 24–48 h post-electroporation.

Microscopic Technique
Live cell microscopy

Cell Type(s)
U2OS 2-6-3 cells