pSEVA3b61
(Plasmid
#58310)
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Purpose(Empty Backbone) Shuttle vector: pUC ori, pTHT15 ori for use in E. coli/B. subtilis. Cannot modify orientation of cat gene; promoter too weak in E. coli, probably transcribed via repB promoter in bicistronic manner.
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Depositing Lab
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Publication
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Sequence Information
Ordering
Item | Catalog # | Description | Quantity | Price (USD) | |
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Plasmid | 58310 | Standard format: Plasmid sent in bacteria as agar stab | 1 | $85 |
Backbone
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Vector backboneSEVA
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Vector typeBacterial Expression, Synthetic Biology
Growth in Bacteria
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Bacterial Resistance(s)Chloramphenicol, 6 μg/mL
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Growth Temperature37°C
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Growth Strain(s)DH10B
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Growth instructionsIn E. coli, use chloramphenicol at 6 µg/mL (gene expression of cat not very strong). In B. subtilis, the same concentration is sufficient for selection.
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Copy numberHigh Copy
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
Despite multiple attempts, we could never clone the cat gene in pSEVA3b61 in the opposite orientation. We concluded that the cat promoter was not sufficient for chloramphenicol resistance in E. coli; rather, the gene was relying on read-through from the upstream repB promoter. We tried several variants extending the cmR promoter region without success. It is important to note that the cat gene works just fine in B. subtilis in either orientation. We found the following paper useful for background reading on this particular cat gene: Olson, D. G., & Lynd, L. R. (2012). Computational design and characterization of a temperature-sensitive plasmid replicon for gram positive thermophiles. Journal of Biological Engineering, 6(1), 5. doi:10.1186/1754-1611-6-5
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:
pSEVA3b61 was a gift from Tom Ellis (Addgene plasmid # 58310 ; http://n2t.net/addgene:58310 ; RRID:Addgene_58310) -
For your References section:
GeneGuard: a Modular Plasmid System Designed for Biosafety. Wright O, Delmans M, Stan GB, Ellis T. ACS Synth Biol. 2014 May 13. 10.1021/sb500234s PubMed 24847673