pX330-Flag-SpCas9-HF1 (without sgRNA)
PurposeExpression plasmid for human codon-optimized high-fidelity SpCas9-HF1, px330-like backbone (without U6-sgRNA coding sequence)
|Item||Catalog #||Description||Quantity||Price (USD)|
|Plasmid||92102||Standard format: Plasmid sent in bacteria as agar stab||1||$65|
This material is available to academics and nonprofits only.
Vector backbonepX330-like (without U6-sgRNA coding sequence)
- Total vector size (bp) 8077
Vector typeMammalian Expression, CRISPR
Growth in Bacteria
Growth Strain(s)NEB Stable
Copy numberHigh Copy
Gene/Insert name3xFlag-NLS-Streptococcus pyogenes Cas9-HF1-NLS
Insert Size (bp)4272
MutationN497A, R661A, Q695A, Q926A
- Promoter Cbh
/ Fusion Proteins
- 3xFLAG (N terminal on insert)
- NLS (N terminal on insert)
- NLS (C terminal on insert)
- Cloning method Unknown
- 5′ sequencing primer GGAAGCAGCGGACCTTCGAC
- 3′ sequencing primer GGAAAGGACAGTGGGAGTGG (Common Sequencing Primers)
Terms and Licenses
pCbh-3xFLAG-NLS-SpCas9-HF1-NLS (without U6-sgRNA coding sequence).
SpCas9-HF1 subcloned in the same plasmid backbone and tailored to identically possess the same NLS and FLAG tags at their termini like the eSpCas9 (without U6-sgRNA coding sequence) (Addgene #92354).
The expression level of SpCas9-HF1 is higher from this than from the original plasmid (VP12; Addgene #72247).
The lack of sgRNA expression cassette allows for easy testing of various SpCas9 variants with the same sgRNA expressed from a separate plasmid (e.g. pmCherry_gRNA, Addgene# #80457).
For plasmid usage and detailed informations, please see the associated publication.
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:pX330-Flag-SpCas9-HF1 (without sgRNA) was a gift from Ervin Welker (Addgene plasmid # 92102 ; http://n2t.net/addgene:92102 ; RRID:Addgene_92102)
For your References section:Crossing enhanced and high fidelity SpCas9 nucleases to optimize specificity and cleavage. Kulcsar PI, Talas A, Huszar K, Ligeti Z, Toth E, Weinhardt N, Fodor E, Welker E. Genome Biol. 2017 Oct 6;18(1):190. doi: 10.1186/s13059-017-1318-8. 10.1186/s13059-017-1318-8 [pii] PubMed 28985763