401 X20 FUSE5-based phage display library 20 5.0e8 Baruch Stern (Department of Cell Research and Immunology, George S Wise Faculty of Life Science, Tel-Aviv University, Israel) Completely random NNK Linear The library was constructed by cloning a 60-nucleotide-long random sequence (correspondmg to a 20-amino acid random sequence) into the pa 402 f88-4/Cys6 phage display library (X4CX6CX4) 16 G. Smith (University of Missouri, Columbia, MO) Semi-random NNM Circular The f88-cys6 is a 16 amino acid sequence randomized for every residue except for the locations of two cysteines and containing more than 2.7e8 phage. 403 T7 PXTGTWX8G phage display library 8 Watanabe H and Yamasaki K (The Biomedical Research Institute, National Institute of Advanced Industrial Science and Technology) Semi-random NNK Linear The library was generated by elongating a segment containing an eight-residue randomized region and a glycine residue Xaa8-Gly at the C terminus of a chignolin-derived segment termed CLN. Here, the Xaa was encoded by a degenerate codon NNK. The gene fragments were synthesized by overlap extension polymerase chain reaction (PCR), digested by restriction enzymes EcoRI/HindIII, and then ligated into the C-terminal part of a T7 phage coat protein gene 10 (g10) of the T7Select 10 -3b vector (Novagen). 404 X28 and X28 M13 phage display library pool 28 Completely random NNK Linear Two libraries displayed 28 amino acid-long random peptides as fusions to the major capsid protein VIII of M13. 405 Twelve different M13 phage libraries pool Completely random NNK Linear 406 X20 phage display library 20 Pillutla RC and Hsiao K (DGI BioTechnologies, Inc, Edison NJ, USA) Completely random DNA fragments coding for peptides containing 20 random amino acids were generated by a PCR approach using synthetic oligonucleotides. Peptides are expressed on the capsid protein pIII of the phage at low copy number (1–2 peptides/phage). 407 X40 phage display library 40 Pillutla RC and Hsiao K (DGI BioTechnologies, Inc, Edison NJ, USA) Completely random DNA fragments coding for peptides containing 40 random amino acids were generated by a PCR approach using synthetic oligonucleotides. Peptides are expressed on the capsid protein pIII of the phage at low copy number (1–2 peptides/phage). 408 fUSE5 X5WYA[WF]SPX4 phage dsiplay library 15 Landon LA (University of Missouri, Columbia, Missouri, USA) Semi-random NNK Linear The plus-strand sequence was 5'-TCG GCC GAC GGG GCC (NNK)5 TGG TAT GCG TGG TCC CCG (NNK)4 GGG GCC TCT GGG GCC GAA AGT-3'. The library is based on FUSE5 vector. Random peptides were express on protein III. The resulting peptides contained the six-amino-acid TF antigen–binding consensus sequence (W-Y-A-W/F-S-P) in the conserved central part of the peptide and terminal regions of random amino acid sequence. The large size of the input library (1.0e11 bacteriophage particles) indicated that a large percentage of the possible 15-amino-acid peptides were represented in the library. 409 CX8C T7 phage display library 8 5.0e8 Pilch J (Burnham Institute for Medical Research, USA) Completely random NNK Circular The cyclic peptide library with the general structure of CX8C was based on a T7 10–3b phage vector. 410 T9 M13 phage display library (X22) 22 2.0e8 Brian K. Kay Completely random NNK Linear The library contains 2e8 unique clones expressing 22-amino acid-long random peptides fused to the amino terminus of M13 protein Ⅲ. 411 T12 M13 phage display library (X36) 36 2.0e8 Brian K. Kay Completely random NNK Linear The library contains 2e8 unique clones expressing 36-amino acid-long random peptides fused to the amino terminus of M13 protein Ⅲ. 412 X6PX2PX6 M13 phage display library 16 3.0e8 Brian K. Kay Semi-random NNK Linear Peptides were expressed as N-terminal fusions to the mature protein Ⅲ of Bacteriophage M13. 413 X12 M13 phage display ibrary 12 Brian K. Kay Completely random NNK Linear The library was based on M13 phage. 414 f3-7mer fUSE5-based phage display library 7 G. Smith (University of Missouri, Columbia, MO) Completely random NNK Linear Library f3-7mer is based on vector fUSE5, which displys its guest peptides on the minor coat protein pIII at one tip of the virion. 415 X8CX8 f88-based phage display library 17 2.5e8 J. K. Scott (Simon Fraser University, Burnaby, Canada) Semi-random NNK Linear The library is based on vector f88, which displays its guest peptides on the coat protein pVIII. 416 X4CX4CX4 f88-4-based phage display library 14 G. Smith (University of Missouri, Columbia, MO) Semi-random NNK Circular The peptides were displayed as fusions with the pⅧ coat protein of the f88-4 phage display vector. 417 pComb3.5 XSX5 phage display library 7 Barbas CF 3rd Semi-random NNK Linear The murine Cys2-His2 zinc finger protein Zif268 was used for construction of phage-display libraries. Primer pairs Aseq (5'-GTCCATAAGATTAGCGGATCC-3') and Zfl6rb [5'-CTGGCCTGTGTGGATGCGGATATG(MNN)5CGAMNNAGAAAAGCGGCGATCGCAGGA-3', where N is A, T, G, or C and M is A or C], and Bseq (5'-GTGAGCGAGGAAGCGGAAGAG-3') and Zflf (5'-CATATCCGCATCCACACAGGCCAG-3') were used to amplify fragments of the Zif268 gene using plasmid pAra-Zif268 as a template. The two PCR fragments were used as templates for overlap extension, and the library was constructed in pComb3.5. 418 pComb3.5 X6 phage display library 6 Barbas CF 3rd Completely random NNK Linear The murine Cys2-His2 zinc finger protein Zif268 was used for construction of phage-display libraries. Primer pairs Aseq (5'-GTCCATAAGATTAGCGGATCC-3') and Zff2r6f [5'-CAGTGTCGAATATGCATGCGTAACTTC(NNK)6ACCACCCACATCCGCACCCAC-3', where N is A, T, G, or C and K is G or T], and Bseq (5'-GTGAGCGAGGAAGCGGAAGAG-3') and Zfnsilb (5'-CATGCATATTCGACACTGGAA-3') were used to amplify fragments of the Zif268 gene using plasmid pAra-Zif268 as a template. The two PCR fragments were used as templates for overlap extension, and the library was constructed in pComb3.5. 419 pComb3H SX4LX2 phage display library 8 4.4e9 Barbas CF 3rd Semi-random NNK Linear The zinc finger library was based on vector pComb3H. It involved randomization of residues within the a-helix of finger 2 of C7, a variant of Zif268. The NNK library was constructed by randomization of positions -1, 1, 2, 3, 5, and 6 by using a condon doping strategy that allows for all amino acid combinations within 32 condons. The position -2 is somewhat artifactual; the NNK library had this residue fixed as serine. Also it contained an invariant leucine at position 4. 420 pComb3H X5LX2 phage display library 8 3.5e9 Semi-random Linear The zinc finger library is based on vector pComb3H. It involved randomization of residues within the a-helix of finger 2 of C7, a variant of Zif268. The library was constructed by randomization of positions -2, -1, 1, 2, 3, 5, and 6, which precludes Tyr, Phe, Cys, and all stop condons in its 24-codon set and contained an invariant leucine at position 4. 421 X10 phagemid-based fd phage display library 10 6.0e9 Completely random NNK Linear Random peptides were fused to the NH2-terminus of pⅧ. 422 X12 phagemid-based fd phage display library 12 2.0e8 Completely random NNK Linear Random peptides were fused to the NH2-terminus of pⅧ. 423 CX8C phagemid-based fd phage display library 8 2.0e9 Completely random NNK Circular Random peptides were fused to the NH2-terminus of pⅧ. 424 pZif12 X15 phage display library 15 Carl O. Pabo (Massachusetts Institute of Technology, Department of Biology, Cambridge, USA) Completely random Linear Phagemid vectors used in the selections were created from pZif12 by restoring the reading frame between the Zif12-coding region and gene III and by introducing convenient restriction sites at the start of Zif12. The library containing randomized 15-residue peptides was constructed by cassette mutagenesis, using NN(G/C/T) randomized codons. The complete fusion protein used for phage display contained a PelB signal sequence; a short leader peptide (NH2-EPRAQNS); the random peptide; residues 4–60 of Zif268; a linker that includes an amber codon; and residues 23–424 of M13 gene III product. 425 XnCXnCXn and Xn M13 phage display library pool ~4.7e11 Gregory A. Weiss (University of California, Irvine, California) Semi-random NNK Linear The phage display library pool was composed of 15 different scaffolds that feature different arrangements of conformation biasing disulfides and randomized residues. XnCXnCXn and Xn type libraries fused to the P8 major coat protein of M13 phage were constructed.