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PurposeCMV and T7 promoter expression plasmid for human codon optimized SpCas9 with a c-terminal bi-partite NLS, 3x flag tag, and P2A-EGFP
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Depositing Lab
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Depositing OrganizationMassachusetts General Hospital
Located in United States of America -
Citations
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Sequence Information
Ordering
| Item | Catalog # | Description | Quantity | Price (USD) | |
|---|---|---|---|---|---|
| Plasmid | 139987 | Standard format: Plasmid sent in bacteria as agar stab | 1 | $94 | |
| DNA | 139987-D50 | 50 μg of DNA in Tris buffer | $495 | ||
| DNA | 139987-D100 | 100 μg of DNA in Tris buffer | $585 | ||
Backbone
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Vector backboneJDS246
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Backbone manufacturerKeith Joung (Addgene Plasmid 43861)
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Vector typeMammalian Expression, CRISPR ; in vitro transcription; T7 promoter
Growth in Bacteria
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Bacterial Resistance(s)Ampicillin, 100 μg/mL
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Growth Temperature37°C
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Growth Strain(s)DH5alpha
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Copy numberHigh Copy
Gene/Insert
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Gene/Insert namehuman codon optimized SpCas9 with BPNLS-3xFLAG-P2A-EGFP
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Alt nameRTW3027
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SpeciesSynthetic
- Promoter CMV and T7
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Tag
/ Fusion Protein
- BPNLS-3xFLAG-P2A-EGFP (C terminal on insert)
Cloning Information
- Cloning method Gibson Cloning
- 5′ sequencing primer oBK6928-CCAAGTCTCCACCCCATTGACG
- 3′ sequencing primer oBK219-GGGAGTGGCACCTTCCAGGGTC
- (Common Sequencing Primers)
Resource Information
Terms and Licenses
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Academic/Nonprofit Terms
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Industry Terms
- Not Available to Industry
Trademarks:
- Zeocin® is an InvivoGen trademark.
DNA (Catalog # 139987-D50)
Purpose
Ready-to-use, high-purity DNA prep. DNA aliquots are suitable for use in mammalian cell transfections and other molecular biology applications.
Delivery
- Amount 50 μg
- Concentration 1 μg/μL
- Pricing $495 USD
- Storage 4 ℃ (short-term) or -20 ℃ (long-term)
Terms and Licenses
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Academic/Nonprofit Terms
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Industry Terms
- Not Available to Industry
Quality Control
Every DNA prep is verified by next-generation sequencing, confirming plasmid identity, sequence integrity, and absence of DNA contaminants. Concentration is confirmed to be within ±2.5% of the stated concentration.
Our production process has been validated to consistently yield endotoxin levels below 1.0 EU/μg. Individual lots are not tested for endotoxin.
DNA (Catalog # 139987-D100)
Purpose
Ready-to-use, high-purity DNA prep. DNA aliquots are suitable for use in mammalian cell transfections and other molecular biology applications.
Delivery
- Amount 100 μg
- Concentration 1 μg/μL
- Pricing $585 USD
- Storage 4 ℃ (short-term) or -20 ℃ (long-term)
Terms and Licenses
-
Academic/Nonprofit Terms
-
Industry Terms
- Not Available to Industry
Quality Control
Every DNA prep is verified by next-generation sequencing, confirming plasmid identity, sequence integrity, and absence of DNA contaminants. Concentration is confirmed to be within ±2.5% of the stated concentration.
Our production process has been validated to consistently yield endotoxin levels below 1.0 EU/μg. Individual lots are not tested for endotoxin.
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.
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For your Materials & Methods section:
pCMV-T7-SpCas9-P2A-EGFP (RTW3027) was a gift from Benjamin Kleinstiver (Addgene plasmid # 139987 ; http://n2t.net/addgene:139987 ; RRID:Addgene_139987) -
For your References section:
Unconstrained genome targeting with near-PAMless engineered CRISPR-Cas9 variants. Walton RT, Christie KA, Whittaker MN, Kleinstiver BP. Science. 2020 Apr 17;368(6488):290-296. doi: 10.1126/science.aba8853. Epub 2020 Mar 26. 10.1126/science.aba8853 PubMed 32217751
