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PurposeBackbone of pComb3X containing TT Fab (light and heavy chains) against tetanus toxin, used as phage display/expression control and PCR template for human antibody constant regions (kappa and CH1)
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Depositing Lab
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Sequence Information
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Sequences (4) — Accept Affinity Reagent Sequence Policy
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Ordering
Item | Catalog # | Description | Quantity | Price (USD) | |
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Plasmid | 63891 | Standard format: Plasmid sent in bacteria as agar stab | 1 | $85 |
Backbone
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Vector backbonepComb3X
- Backbone size w/o insert (bp) 3300
- Total vector size (bp) 4700
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Vector typeBacterial Expression ; phage display
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)XL1 Blue
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Copy numberHigh Copy
Gene/Insert
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Gene/Insert nameTT Fab
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Alt namehuman Fab to tetanus toxin
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SpeciesH. sapiens (human)
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Insert Size (bp)1400
- Promoter lacZ
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Tags
/ Fusion Proteins
- 6xHis (C terminal on backbone)
- HA tag (C terminal on backbone)
- geneIII (C terminal on backbone)
Cloning Information
- Cloning method Restriction Enzyme
- 5′ cloning site SfiI (not destroyed)
- 3′ cloning site SfiI (not destroyed)
- 5′ sequencing primer AAG ACA GCT ATC GCG ATT GCA G
- 3′ sequencing primer GCC CCC TTA TTA GCG TTT GCC ATC (Common Sequencing Primers)
Resource Information
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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.
Depositor Comments
pComb3X is the newest of the pComb vectors. Improvements over pComb3 include increased stability and introduction of an asymmetric SfiI cassette for directional cloning of full Fab, scFv, peptide and other protein for phage display. 6xHis and HA tags allow for purification and detection. An amber stop codon was introduced to turn-off expression of the pIII fusion protein by switching to a non-supressor strain of E. coli allowing production of soluble protein without subcloning. Alternatively, the gene for phage protein pIII can be removed by SpeI/NheI digest. The “TT” refers to the human Fab to tetanus toxin which can be used as a template to amplify the kappa light chain constant region and Fd for construction of chimeric antibody libraries as described in:
Barbas, C. F., III; Burton, D. R.; Scott, J.K., Silverman, G.J. Eds. (2001) Phage Display: A Laboratory Manual; Cold Spring Harbor Laboratory Press: Cold Spring Harbor, New York
It can also be used as a phage display/protein expression control.
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:
pComb3XTT was a gift from Carlos Barbas (Addgene plasmid # 63891 ; http://n2t.net/addgene:63891 ; RRID:Addgene_63891) -
For your References section:
Methods for the generation of chicken monoclonal antibody fragments by phage display. Andris-Widhopf J, Rader C, Steinberger P, Fuller R, Barbas CF 3rd. J Immunol Methods. 2000 Aug 28;242(1-2):159-81. 10.1016/s0022-1759(00)00221-0 PubMed 10986398