PurposeLentiCRISPR v2 was modified into an all-in-one dox inducible system. The addition of doxycycline induces Cas9-2A-eGFP. The U6 promoter drives constitutive sgRNA expression.
|Item||Catalog #||Description||Quantity||Price (USD)|
|Plasmid||87360||Standard format: Plasmid sent in bacteria as agar stab||1||$65|
This material is available to academics and nonprofits only.
Vector backbonelentiCRISPR v2
Backbone manufacturerFeng Zhang lab
- Backbone size w/o insert (bp) 16733
Modifications to backboneLentiCRISPR v2 was modified into an all-in-one inducible system, doxycycline induces Cas9-P2A-GFP expression. sgRNA and puromycin expression are driven by constitutive promoters. The following modifications were made to lentiCRISPR v2: the EF1 promoter was removed and replaced with a tight TRE promoter; P2A-puromcycin was removed and replaced with T2A-eGFP; and a EF1 promoter-puromycin-P2A-rtTA cassette was added.
Vector typeMammalian Expression, Lentiviral, CRISPR ; Doxycycline inducible; eGFP reporter
Growth in Bacteria
Growth Strain(s)NEB Stable
- Promoter Tight TRE promoter
- Cloning method Unknown
- 5′ sequencing primer LNCX [5'-AGCTCGTTTAGTGAACCGTCAGATC-3']
- 3′ sequencing primer EF1a-R [5'CACGGCGACTACTGCACTTA-3'] (Common Sequencing Primers)
Clone sgRNAs into TLCV2 using the same protocol provided with lentiCRISPR v2 (Plasmid #52961, protocol listed under the RESOURCE INFORMATION section). After you have inserted your sgRNA, you should sequence this region with one of the following primers: hU6-F (5'-GAGGGCCTATTTCCCATGATT-3') or LKO.1 5’(5'- GACTATCATATGCTTACCGT-3').
These plasmids were created by your colleagues. Please acknowledge the Principal Investigator, cite the article in which the plasmids were described, and include Addgene in the Materials and Methods of your future publications.
For your Materials & Methods section:TLCV2 was a gift from Adam Karpf (Addgene plasmid # 87360 ; http://n2t.net/addgene:87360 ; RRID:Addgene_87360)
For your References section:Pan-Cancer Analyses Reveal Genomic Features of FOXM1 Overexpression in Cancer. Barger CJ, Branick C, Chee L, Karpf AR. Cancers (Basel). 2019 Feb 21;11(2). pii: cancers11020251. doi: 10.3390/cancers11020251. 10.3390/cancers11020251 PubMed 30795624