Checking Transformation Efficiency of Chemically Competent Cells

SUPPLIES:

Equipment:

• Shaking Incubator or Shaking Platform and 1L flask clamps
• 42°C Heating Bath or Block
• 37°C Incubator
• Colony Counter

Consumables:

• Agar plates with appropriate antibiotic

Buffers and Solutions:

• Competent cells prepared using the Inoue method
• 10pg / μL solution of a standard plasmid (e.g. pUC19)
• SOC Medium

PROTOCOL:

1. ) Transform 10pg of a standard plasmid (e.g. pUC19) in 50μL of cells using the standard transformation protocol.
2. ) Plate 50uL of transformed cells in triplicate on appropriate antibiotic
3. ) Grow plates overnight, and count colonies the next morning
4. ) Average colony counts for the three plates; efficiency = (# colonies) x 10⁶ cfu / μg
5. ) Expected efficiency for the Inoue protocol is 1-3 x 10⁸ colonies / μg of plasmid

-- Main.KateElston - 25 Oct 2018