...A549 cells stably expressing Cas9. A549 cells were transduced with lentiCas9-Blast 1 and then selected with...Figure 2: Cas9 expression in monoclonal cell lines generated from A549 cells transduced with lentiCas9-Blast... express Cas9 and are included as a control (WT). Vinculin is a loading control. 1 lentiCas9-Blast was...additional 3 weeks under blasticidin selection. Anti-Cas9 Western blotting was performed on whole-cell extracts...was a gift from Feng Zhang (Addgene plasmid #52962 ) and is described in Improved vectors and genome-wide...

...Storing and Handling Addgene Plasmids What to do after you receive your plasmids from Addgene Instructions...Protocol Diagnostic Restriction Digest Verify a plasmid using restriction enzymes Diagnostic Restriction...Instructions for Storing and Handling Plasmids Genomic Deletions with CRISPR Generate genomic deletions in mammalian...mammalian cells lines using CRISPR/Cas9 Genomic Deletions with CRISPR Protocol Over-Agar Antibiotic Plating...Transformations We walk you through a standard plasmid transformation protocol and offer some tips and...Center How to Deposit Your guide to depositing plasmids with Addgene Depositing Page Career Videos (Link...

...A549 cells stably expressing Cas9. A549 cells were transduced with lentiCas9-Blast and then selected with...single cells for 13 days. lentiCas9-Blast was a gift from Feng Zhang (Addgene plasmid #52962 ) and is described...

... cm dish). Using transfer plasmid pHAGE TRE dCas9-KRAB (total μg of plasmid DNA 27.8 μg), this would be... 0.72 pmol Transfer Plasmid* 1.64 pmol OptiPro SFM to total volume 500 μL *Plasmid concentrations and ... each transfer plasmid. Pro-Tip Endotoxins can inhibit transfection; therefore, plasmid DNA purification...endotoxin removal step. For high quality plasmid DNA, the plasmids should also be propagated in an endonuclease...~20 h. Prepare a mixture of the 3 transfection plasmids into OptiPro SFM: Reagent Amount per 10 cm dish...is prepared, transfect cells with a fluorescent plasmid using a variety of ratios. Check the cells 1-2 ...cells were transfected with the GFP-expression plasmid pRosetta using μg total DNA to μg PEI ratios of...