Duke University/24 June 2009

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June 24, 2009

 * E: Gel to check PCR
 * R: Correct band (faint)
 * C: Do transformation with competent cells

Transformation with High Efficiency GC5 Competent Cells
 * 1) Remove competent cells from -70°C and place on ice. Thaw for 5-10 min.
 * 2) Gently mix cells by tapping tube.
 * 3) Add 1-50 ng DNA (1 ul control) into 50 ul cells. Swirl pipette tip while dispensing DNA. Gently tap tube to mix.
 * 4) Place tubes on ice for 30 min.
 * 5) Heat-shock cells for 45 sec in 42°C (water) bath. Do not shake!
 * 6) Add 450 ul RT SOC Medium to each transformation reaction.
 * 7) Incubate at 37°C for 1 hr with shaking at 225-250 rpm.
 * 8) Spread on LB agar plates containing appropriate antibiotic.
 * 9) Incubate plates at 37°C overnight (12-16 hrs).