lentiCRISPR v2

Protocol tips

Upstream tips	Protocol tips	Downstream tips
	There is no need to perform a negative control golden-gate reaction (without insert) as it will always contain colonies so not a good indicator of cloning success.

Protocol tips
There is no need to perform a negative control golden-gate reaction (without insert) as it will always contain colonies so not a good indicator of cloning success.

Publication protocol

Lentiviral vector production- The lentiviral CRISPR/Cas9 mediated miR-21 gene editing vectors were constructed by annealing four gRNA oligonucleotide pairs and subcloning them in the BsmII sites of lentiviral vector pLenti CRISPR V2. A control vector was constructed by inserting EGFP gRNA sequences into pLenti CRISPR V2 lentiviral vector. All gRNA sequences were selected from the Human GeCKOv2 CRISPR knockout pooled library 10 and primers for detecting miR-21 mutations in surveyor mutation assay are listed in Table ​Table1.1. Lentivirus were produced by packaging in 293FT cells as we published previously 11. Stable cell lines were generated by transducing the SKOV3 and OVCAR3 cells with the lentiviral CRISPR/Cas9 miR-21 gene editing vectors and selected with 5 μg/ml puromycin.

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Papers

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Manufacturer protocol

Download the product protocol from Addgene for lentiCRISPR v2 below.

Download manufacturer protocol

Videos

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