Plating Protocol for NEB PCR Cloning Kit

  1. Mix cells thoroughly by flicking or inversion and spread 50 μl of the 1 ml outgrowth of a 1:10 dilution onto 37°C pre-warmed agar plates containing 100 μg/ml ampicillin. For 15  minute ligations, it is especially important to spread both volumes of outgrowth as the selection for cloned inserts is strongest with plating densities < 400 colonies per plate.

  2. Invert plate and incubate overnight at 37°C or for 24 hours at 30°C. Do not use room temperature growth as the slow growth rate will interfere with selection of constructs wi,th inserts.