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PurposeCMV and T7 promoter expression plasmid for human codon optimized SpRY(A61R/L1111R/D1135L/S1136W/G1218K/E1219Q/N1317R/A1322R/R1333P/R1335Q/T1337R) with a c-terminal bi-partite NLS, 3x flag tag, and P2A-EGFP
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Depositing Lab
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Sequence Information
Ordering
Item | Catalog # | Description | Quantity | Price (USD) | |
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Plasmid | 139989 | Standard format: Plasmid sent in bacteria as agar stab | 1 | $85 |
Backbone
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Vector backboneRTW3027
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Backbone manufacturerKleinstiver Lab (Addgene ID 139987)
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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 variant named SpRY with BPNLS-3xFLAG-P2A-EGFP
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Alt nameRTW4830
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SpeciesSynthetic
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MutationSpRY=A61R/L1111R/D1135L/S1136W/G1218K/E1219Q/N1317R/A1322R/R1333P/R1335Q/T1337R
- 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
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Supplemental Documents
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Articles Citing this Plasmid
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.
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-SpRY-P2A-EGFP (RTW4830) was a gift from Benjamin Kleinstiver (Addgene plasmid # 139989 ; http://n2t.net/addgene:139989 ; RRID:Addgene_139989) -
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