pET24a_UDP_Xase
(Plasmid
#73158)
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PurposeExpresses in E. coli, S. pneumoniae TIGR4 UDP-Xase SP_1669
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Depositing Lab
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Sequence Information
Full plasmid sequence is not available for this item.
Ordering
Item | Catalog # | Description | Quantity | Price (USD) | |
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Plasmid | 73158 | Standard format: Plasmid sent in bacteria as agar stab | 1 | $85 |
Backbone
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Vector backbonepET24
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Backbone manufacturerNovagen
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Vector typeBacterial Expression
Growth in Bacteria
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Bacterial Resistance(s)Kanamycin, 50 μg/mL
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Growth Temperature37°C
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Growth Strain(s)DH5alpha
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Growth instructionsFor protein production, use BL21 DE3 cells. Cells were grown at 37°C in LB media. When an OD600 of 0.9 was reached, 1 mM IPTG was added to induce expression. After overnight growth at 22°C, the cells were centrifuged, washed in 50 mM Tris pH 7.5, 1 mM EDTA, and 0.1 mM DTT (TED buffer), reharvested, and stored at −80°C.
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Copy numberLow Copy
Gene/Insert
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Gene/Insert namepET24a
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Alt nameA0A0H2URA0
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Alt nameNC_003028.3
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Alt nameSp_1699
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SpeciesStreptococcus pneumoniae serotype 4 (strain ATCC BAA-334 / TIGR4
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GenBank IDNC_003028.3
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
The over-expressed protein was released upon thawing of the
cells. The cell extract was adjusted to 33% ammonium sulfate
saturation and allowed to precipitate for 30 minutes at 4uC.
Precipitated protein was harvested by centrifugation at 4uC for 30minutes at 172006g on a GSA rotor (Sorvall). TED buffer was
used to resuspend the precipitated protein. The remaining soluble
protein was brought up to 50% ammonium sulfate saturation and
the above procedure was repeated to harvest and resuspend
precipitated protein. The supernatant after this step was brought
up to 30% glycerol (v/v) and loaded onto a Sephadex, G-100
(2.5650 cm) gel filtration column equilibrated with TED buffer
containing 100 mM NaCl.
THe structure has been determined by X-ray crystallography and deposited in the PDB 4HFQ
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:
pET24a_UDP_Xase was a gift from Sandra Gabelli (Addgene plasmid # 73158 ; http://n2t.net/addgene:73158 ; RRID:Addgene_73158) -
For your References section:
A UDP-X diphosphatase from Streptococcus pneumoniae hydrolyzes precursors of peptidoglycan biosynthesis. Duong-Ly KC, Woo HN, Dunn CA, Xu W, Babic A, Bessman MJ, Amzel LM, Gabelli SB. PLoS One. 2013 May 15;8(5):e64241. doi: 10.1371/journal.pone.0064241. Print 2013. PONE-D-13-06764 [pii] PubMed 23691178