pADL-22c

PD0110

Price (USD): $697.00
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pADL-22c

Technical Description

pADL™-22c 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 HA 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.

 

pADL-22 Western blot

Figure 1. Reactivity of HA tag. Western blot analysis of periplasmic extracts prepared from varied phagemids containing the HyHEL-10 scFv insert. Only the scFv derived from pADL-22 exhibits a strong reactivity with a HA tag antibody (Left Panel; Lane A: pADL-20, Lane B: pADL-22, Lane C: pADL-23). Binding analysis of HyHEL-10 scFv by ELISA using a HA tag antibody as a secondary antibody (Right Panel; PERI: periplasmic extract, SUP bacterial supernatant, LYZ: lyzozyme, BSA: bovine serum albumin). Note: Expression of pADL-22c is 5 to 10 times higher than the parent vector pADL-22.

Applications

  • Phage display at the N-terminal side of gene III protein of the filamentous bacteriophage.

  • Free scFv or Fab expression.

For research use only; not intended for any animal or human therapeutic or diagnostic use.

Specifications

General Characteristics:

Plasmid Size: 3957
Promoter: lac promoter
Leader Peptide: PelB
Cloning Site: double-SfiI/BglI, NotI-SpeI
Purification: HIS tag
Detection: HA tag
Fusion Protein: full length gene III protein and conditional Amber stop codon
Selection: ampicillin
Replication: oriF1, pMB1

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.

Quality Control & Certification of Analysis

Product Size:

Digestion by EcoRI and NheI generates 2 fragments of 2.6 kb and 1.4 kb.

Product Sequence:

Complete vector sequence is analyzed.

Certification:

Product meets all specifications.

pADL-22c map

 

pADL-22c cloning site

 

Citations

  1. Yuan TZ, Garg P, Wang L, Willis JR, Kwan E, Hernandez AGL, Tuscano E, Sever EN, Keane E, Soto C, Mucker EM, Fouch ME, Davidson E, Doranz BJ, Kailasan S, Aman MJ, Li H, Hooper JW, Saphire EO, Crowe JE, Liu Q, Axelrod F, Sato AK (2022). Rapid discovery of diverse neutralizing SARS-CoV-2 antibodies from large-scale synthetic phage libraries. MAbs. 2022;14(1):2002236.

  2. Ye G, Pan R, Bu F, Zheng J, Mendoza A, Wen W, Du L, Spiller B, Wadzinski BE, Liu B, Perlman S, Li F (2023). Discovery of Nanosota-2, -3, and -4 as super potent and broad-spectrum therapeutic nanobody candidates against COVID-19. J Virol. 2023;97(11):e0144823.

  3. Kordus SL, Kroh HK, Rodríguez RC, Shrem RA, Peritore-Galve FC, Shupe JA, Wadzinski BE, Lacy DB, Spiller BW (2023). Nanobodies against C. difficile TcdA and TcdB reveal unexpected neutralizing epitopes and provide a toolkit for toxin quantitation in vivo. PLoS Pathog. 2023;19(10):e1011496.

  4. Schlör A, Hirschberg S, Amor GB, Meister TL, Arora P, Pöhlmann S, Hoffmann M, Pfaender S, Eddin OK, Kamhieh-Milz J, Hanack K. (2022). SARS-CoV-2 neutralizing camelid heavy-chain-only antibodies as powerful tools for diagnostic and therapeutic applications. Front Immunol., 14;13:930975.

  5. Yuan T.Z., Garg P., Wang L., Willis J.R., Kwan E., Hernandez A.G.L., Tuscano E., Sever E.N., Keane E., Soto C., Mucker E.M., Fouch M.E., Davidson E., Doranz B.J., Kailasan S., Aman M.J., Li H., Hooper J.W., Saphire E.O., Crowe J.E., Liu Q., Axelrod F., Sato A.K. (2022). Rapid discovery of diverse neutralizing SARS-CoV-2 antibodies from large-scale synthetic phage libraries. MAbs., 14(1):2002236.

  6. Wu Y., Chen Z., Sigworth F.J., Canessa C.M. (2021). Structure and analysis of nanobody binding to the human ASIC1a ion channel. Elife, 10:e67115.

  7. Salem, R., El-Kholy, A.A., Ibrahim, M. (2019). Eight novel single chain antibody fragments recognising VP2 of foot-and-mouth disease virus serotypes A, O, and SAT 2. Virology, 533:145-154.

  8. Nelson, R.S. & Valadon; P. (2017). A universal phage display system for the seamless construction of Fab libraries. J Immunol Methods, 450:41-49.