pADL-23c Phagemid Vector

Product Description

pADL™-23c is a phagemid vector designed for phage display on the N-terminal side of the protein III of the filamentous bacteriophage M13 or equivalent. This vector contains a PelB leader sequence for expression in the periplasm, a double-SfiI cloning site to introduce scFvs or Fab fragments, a HIS tag for purification, a Myc tag for detection and an amber codon located before the full-length copy of the gene III sequence. On non-suppressive bacterial strains, free scFvs or Fab fragments are produced in the periplasm where they can be assayed for binding or purified for further testing. Expression of the fusion is under the control of a lac promoter that has been adjusted at a level comparable to the widely used phagemid pCOMB3.

Figure 1. Binding of Myc tag antibody to HyHEL-10 scFv. Western blot analysis of periplasmic extracts prepared from varied phagemids containing the HyHEL-10 scFv insert. Only the scFv derived from pADL-23 exhibits a strong reactivity with a Myc tag antibody (Lane A: pADL-20, Lane B: pADL-22, Lane C: pADL-23).

Vector Map

Cloning Site

Applications

• Phage display at the N-terminal side of gene III protein of the filamentous bacteriophage.
• Free scFv or Fab expression.

Specifications

General characteristics:

• Plasmid Size: 3960
• Promoter: lac promoter
• Leader Peptide: PelB
• Cloning Sites: double-BglI/SfiI, NotI-SpeI
• Purification: HIS tag
• Detection: Myc tag
• Fusion Protein: full length gene III protein and conditional Amber stop codon
• Selection: ampicillin
• Replication: oriF1, pBM1

Physical Characteristics:

• Concentration: 0.5 µg/ µl
• Product Size: 10 µg
• Buffer: DNA Conservation Buffer (Tris/HCL 5 mM, EDTA 0.1 mM, pH 8.5, sterile)
• Storage Temperature: -20°C

Related Products

As helper phage for the preparation of virions we recommend Interference-Resistant Helper Phage CM13.

Citations

1. Melia C.E., Bolla J.R., Katharios-Lanwermeyer S., Mihaylov D.B., Hoffmann P.C., Huo J., Wozny M.R., Elfari L.M., Böhning J., Morgan A.N., Hitchman C.J., Owens R.J., Robinson C.V., O'Toole G.A., Bharat T.A.M. ()2021). Architecture of cell-cell junctions in situ reveals a mechanism for bacterial biofilm inhibition. Proc Natl Acad Sci U S A., 118(31):e2109940118.

2. Gomez-Castillo L., Watanabe K., Jiang H., Kang S., Gu L. (2020). Creating Highly Specific Chemically Induced Protein Dimerization Systems by Stepwise Phage Selection of a Combinatorial Single-Domain Antibody Library. J Vis Exp. , 14;(155):10.3791/60738.

3. Mendoza-Salazar I., Gómez-Castellano KM., González-González E., Gamboa-Suasnavart R., Rodríguez-Luna SD., Santiago-Casas G., Cortés-Paniagua MI., Pérez-Tapia SM., Almagro JC. (2022). Anti-SARS-CoV-2 Omicron Antibodies Isolated from a SARS-CoV-2 Delta Semi-Immune Phage Display Library. Antibodies (Basel),10;11(1):13.

4. Maeda S., Xu J., N Kadji FM., Clark M.J., Zhao J., Tsutsumi N., Aoki J., Sunahara R.K., Inoue A., Garcia K.C., Kobilka B.K. (2020). Structure and selectivity engineering of the M1 muscarinic receptor toxin complex. Science,369(6500):161-167.

5. Huo J., Le Bas A., Ruza RR. et al. (2020). Neutralizing nanobodies bind SARS-CoV-2 spike RBD and block interaction with ACE2. Nat Struct Mol Biol., 27(9):846-854.

6. Gomez-Castillo L., Watanabe K., Jiang H., Kang S., Gu L. (2020). Creating Highly Specific Chemically Induced Protein Dimerization Systems by Stepwise Phage Selection of a Combinatorial Single-Domain Antibody Library. J. Vis. Exp. (155), e60738.

7. Huen J., Yan Z., Iwashkiw J., Dubey S., Gimenez M.C., Ortiz M.E., Patel S.V., Jones M.D., Riazi A., Terebiznik M., Babaei S., Shahinas D. (2019). A Novel Single Domain Antibody Targeting FliC Flagellin of Salmonella enterica for Effective Inhibition of Host Cell Invasion. Front Microbiol., 10,2665.

8. Hassan K.M.A., Hansen J.D., Herrin BR, Amemiya C.T. (2019). Generation of Lamprey Monoclonal Antibodies (Lampribodies) Using the Phage Display System. Biomolecules, 9(12), 868.

9. Valadon P., Pérez-Tapia S.M., Nelson R.S., Guzmán-Bringas O.U., Arrieta-Oliva H.I., Gómez-Castellano K.M., Pohl M.A., Almagro J.C. (2019). ALTHEA Gold Libraries™: antibody libraries for therapeutic antibody discovery. MAbs, 11(3):516-531.

10. WIPO Patent Application (2017). Vectors for cloning and expression of proteins, methods and applications thereof. WO 2017/175176 A1.

11. WIPO Patent Application (2016). Anti-salmonella Antibodies And Uses Thereof. WO 2016/125089 A1.