The CRISPR vector [GeneArt™ CRISPR Nuclease Vector] contain an OFP reporter but is not encoded with antibiotic resistance gene.
12-24 hours following lipofection, there was no significant orange fluorescence under the fluorescent microscope (OFP has a peak excitation of 548nm, and emission of 560nm).
Only few cells were positively transfected this is why FACS enrichment was not feasible. I have repeated this experiment several times with different ratio of DNA:Lipofectamine 2000 but I was still unable to achieve reasonable transfection efficiency.
Any suggestions on how I may improve my protocol to enhance my transfection for knockdown/knockout efficiency?
Thank you.
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