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pADL-10b
Technical Description
pADL™-10b is a phagemid vector designed for phage display on the filamentous bacteriophage M13 or equivalent. This vector contains a PelB leader sequence followed by a full-length copy of the gene III sequence and a double-SfiI cloning site to introduce foreign polypeptides on the N-terminal side of the gene III protein. Expression of the fusion is under the tight control of a lac promoter. A strong transcriptional terminator upstream from the promoter efficiently represses undesirable expression and prevents promoter leakiness in absence of induction, thus limiting negative selection against clones bearing toxic products. The absence of amber codon before the gene III results in strong display (Fig. 1).
Figure 1. HyHEL-10 scFv Display. Western blot analysis of phage particles derived from M13KO7 (lane 1), pADL-10 alone (lane 2) or pADL-10 with the HyHEL-10 scFv insert in varied stages of IPTG induction (lane 3-11). Blot made with a gene III protein antibody.
Applications
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Phage display at the N-terminal side of gene III protein of the filamentous bacteriophage.
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scFv display.
For research use only; not intended for any animal or human therapeutic or diagnostic use.
Specifications
General Characteristics:
Plasmid Size: 4990
Promoter: lac promoter
Leader Peptide: PelB
Cloning Site: double-BglI/SfiI, NotI-SpeI
Fusion Protein: full length gene III protein
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 BamHI, HindIII ad XbaI generates 3 fragments of 3.6 kb, 0.73 kb and 0.64 kb.
Product Sequence:
Sequence of the entie plasmid is analyzed.
Certification:
Product meets all specifications.
Citations
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Aripov VS, Volkova NV, Ilyichev AA, Shcherbakov DN. (2024). Problems of creating antibody phage libraries and their solutions. Vavilovskii Zhurnal Genet Selektsii. 2024;28(2):249-257.
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Wright KM, DiNapoli SR, Miller MS et al. (2023). Hydrophobic interactions dominate the recognition of a KRAS G12V neoantigen. Nat Commun. 2023 Aug 21;14(1):5063.
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Hampton JT, Cho CD, Coleman DD et al. (2023). An amber-encoding helper phage for more efficient phage display of noncanonical amino acids, Nucleic Acids Res., 2023; 51(13): 6566–6577.
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Hampton JT, Lalonde TJ, Tharp JM et al. (2022). Novel Regioselective Approach to Cyclize Phage-Displayed Peptides in Combination with Epitope-Directed Selection to Identify a Potent Neutralizing Macrocyclic Peptide for SARS-CoV-2. ACS Chem. Biol., 17(10):2911-2922.
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Fang Z., Cao D., Qiu J. (2020). Development and production of nanobodies specifically against green fluorescence protein. Appl. Microbiol. Biotechnol., 104(11):4837-4848.
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Tharp J.M., Trae Hampton J., Reed C.A., Ehnbom A., Chen P.C., Morse J.S., Kurra Y., Pérez L.M., Xu S. & Liu W.r. (2020). An amber obligate active site-directed ligand evolution technique for phage display. Nature Comm., 11(1), 1392.
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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.
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Skora A.D., Douglass J., Hwang M.S., Tam A.J., Blosser R.L., Gabelli S.B., Cao J., Diaz L.A. Jr, Papadopoulos N., Kinzler K.W., Vogelstein B., Zhou S. (2015). Generation of MANAbodies specific to HLA-restricted epitopes encoded by somatically mutated genes. Proc Natl Acad Sci U S A, 112(32), 9967–72.
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US Patent Application (2016). Hiv Antigens And Antibodies. US 2016/0002319 A1.
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WIPO Patent Application (2016). Hla-restricted Epitopes Encoded By Somatically Mutated Genes. WO 2016/154246 A1.