pCA24N-ligase-cTF
(Plasmid
#87743)
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PurposeIPTG-inducible expression of ligase-cTF fusion for protein purification
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Depositing Lab
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Sequence Information
Ordering
Item | Catalog # | Description | Quantity | Price (USD) | |
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Plasmid | 87743 | Standard format: Plasmid sent in bacteria as agar stab | 1 | $85 |
Backbone
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Vector backbonepCA24N
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Backbone manufacturerKitagawa et al. (PMID:16769691)
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Vector typeBacterial Expression
Growth in Bacteria
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Bacterial Resistance(s)Chloramphenicol, 25 μg/mL
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Growth Temperature37°C
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Growth Strain(s)DH5alpha
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Copy numberUnknown
Gene/Insert
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Gene/Insert nameT4 DNA ligase
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Entrez Gene30 (a.k.a. T4p202, lig)
- Promoter T5-lac
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Tags
/ Fusion Proteins
- 6xHis (N terminal on backbone)
- cTF (chimeric transcription factor based on NFAT/NF-kB) (C terminal on insert)
Cloning Information
- Cloning method Unknown
- 5′ sequencing primer pCA24N.for (GATAACAATTTCACACAGAATTCATTAAAGAG)
- 3′ sequencing primer pCA24N.rev2 — 5'-CAAATCCAGATGGAGTTCTGAGG-3' (Common Sequencing Primers)
Resource Information
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Supplemental Documents
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A portion of this plasmid was derived from a plasmid made byPreviously, we PCR amplified the lig gene from pRBL (Ren et al., 1997; PMID: 9305776). The cTF gene was amplified from pSAN101 (de Lumley et al., 2004; PMID: 15178248).
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.
Depositor Comments
For complete cloning information, please see the paper's supplement.
For expression, E. coli DH5a-E cells harbouring each expression vector were grown at 37°C in LB broth containing chloramphenicol (34 µg.ml-1), to OD600 ~ 0.6. IPTG (0.4 mM, final concentration) was added as an inducer, and protein over-expression was at 28°C for 16-18 h.
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:
pCA24N-ligase-cTF was a gift from Wayne Patrick (Addgene plasmid # 87743 ; http://n2t.net/addgene:87743 ; RRID:Addgene_87743) -
For your References section:
Engineered DNA ligases with improved activities in vitro. Wilson RH, Morton SK, Deiderick H, Gerth ML, Paul HA, Gerber I, Patel A, Ellington AD, Hunicke-Smith SP, Patrick WM. Protein Eng Des Sel. 2013 Jul;26(7):471-8. doi: 10.1093/protein/gzt024. Epub 2013 Jun 10. 10.1093/protein/gzt024 PubMed 23754529