This week, we will be performing a DpnI digest on our OE-PCR cloning reaction, then transforming it into chemically competent DH5α E. coli by heat shock, and plating them out on LB ampicillin plates.
DpnI Digest, Ligation, and Transformation of Chemically Competent E. coli
Prepare your Plates
Spread 40 L of 1M IPTG, and 40 L of 40mg/mL evenly on the ampicillin μ μ plate. You need to make sure the entire plate is evenly covered.
Always label your plates with your names, the date, and MBHG. When writing on a plate, always write on the back of the plate, not on the lid. The lid can come off and get lost, the back can’t.
Pipette 25 L of the transformed cells onto the plate and spread in a fanμ shape from the line to the plate edges. When you are finished, seal your plate with Parafilm by stretching the Parafilm around the edge of the closed plate (if you don’t know how to do this, ask a demonstrator), and turn in to the instructor for incubation.
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