pADL-22c Phagemid Vector

Product 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.

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.

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: 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

Related Products

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

Citations

1. 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.

2. 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.

3. 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.

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