AMPK Methods and Protocols

pL-CRISPR.EFS.GFP

AMPKα2

sgRNA Cas9 GFP

plentiCRISPR

AMPKα1

sgRNA Cas9 Puromycin

AMPKα1 KO

AMPKα1 KO

AMPKα2 KO

Virus production

Infection

Puromycin selection

Screening

Validation

Virus production

Infection

GFP selection

Screening

Validation

AMPKα1 sgRNA

AMPKα2 sgRNA

A

Fig. 3Experimental workflow for genome engineering of colon carcinoma Caco2 cells. We used a sequential
procedure to achieve both AMPKα1 and AMPKα2 knockout in single-cell Caco2 clones. (A) PRKAA1-targeting
sgRNAs are cloned into the plentiCRISPR plasmid, allowing a selection based on puromycin resistance.
PRKAA2-targeting sgRNAs are cloned into the pL-CRISPR.EFS.GFP plasmid allowing a selection based on GFP

CRISPR-Cas9 For Human AMPK 183