Surface expression libraries of randomized peptides
OwnerWilliam D. Huseinventor
Life of this patent
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Abstract
A composition of matter comprising a plurality of procaryotic cells containing a diverse population of expressible oligonucleotides operationally linked to expression elements, said expressible oligonucleotides having a desirable bias of random codon sequences.
Claims87
A composition of matter comprising a plurality of cells containing a diverse population of expressible oligonucleotides operationally linked to expression elements, said expressible oligonucleotides having a desirable bias of random codon sequences produced from random combinations of first and second oligonucleotide precursor populations having a desirable bias of random codon sequences.
A kit for the preparation of vectors useful for the expression of a diverse population of random peptides from combined first and second oligonucleotides having a desirable bias of random codon sequences, comprising:
- two vectors:
- a first vector having a cloning site for said first oligonucleotides and a pair of restriction sites for operationally combining first oligonucleotides with second oligonucleotides;
- and a second vector having a cloning site for said second oligonucleotides and a pair of restriction sites complementary to those on said first vector, one or both vectors containing expression elements capable of being operationally linked to said combined first and second oligonucleotides.
A cloning system for expressing random peptides from diverse populations of combined first and second oligonucleotides having a desirable bias of random codon sequences, comprising:
- a set of first vectors having a diverse population of first oligonucleotides having a desirable bias of random codon sequences and a set of second vectors having a diverse population of second oligonucleotides having a desirable bias of random codon sequences, said first and second vectors each having a pair of restriction sites so as to allow the operational combination of first and second oligonucleotides into a contiguous oligonucleotide having a desirable bias of random codon sequences.
A composition of matter comprising a plurality of cells containing a diverse population of expressible oligonucleotides operationally linked to expression elements, said expressible oligonucleotides having a desirable bias of random codon sequences.
A plurality of vectors containing a diverse population of expressible oligonucleotides having a desirable bias of random codon sequences.
A composition of matter, comprising a diverse population of oligonucleotides having a desirable bias of random codon sequences produced from random combinations of two or more oligonucleotide precursor populations having a desirable bias of random codon sequences.
A method of constructing a diverse population of vectors having combined first and second oligonucleotides having a desirable bias of random codon sequences capable of expressing said combined oligonucleotides as random peptides, comprising the steps of:
- (a) operationally linking sequences from a diverse population of first oligonucleotides having a desirable bias of random codon sequences to a first vector;
- (b) operationally linking sequences from a diverse population of second oligonucleotides having a desirable bias of random codon sequences to a second vector;
- and
- (c) combining the vector products of steps (a) and (b) under conditions where said populations of first and second oligonucleotides are joined together into a population of combined vectors capable of being expressed.
A method of selecting a peptide capable of being bound by a ligand binding protein from a population of random peptides, comprising:
- (a) operationally linking a diverse population of first oligonucleotides having a desirable bias of random codon sequences to a first vector;
- (b) operationally linking a diverse population of second oligonucleotides having a desirable bias of random codon sequences to a second vector;
- (c) combining the vector products of steps (a) and (b) under conditions where said populations of first and second oligonucleotides are joined together into a population of combined vectors;
- (d) introducing said population of combined vectors into a compatible host under conditions sufficient for expressing said population of random peptides;
- and
- (e) determining the peptide which binds to said ligand binding protein.
A method for determining the nucleic acid sequence encoding a peptide capable of being bound by a ligand binding protein which is selected from a population of random peptides, comprising:
- (a) operationally linking a diverse population of first oligonucleotides having a desirable bias of random codon sequences to a first vector;
- (b) operationally linking a diverse population of second oligonucleotides having a desirable bias of random codon sequences to a second vector;
- (c) combining the vector products of steps (a) and (b) under conditions where said populations of first and second oligonucleotides are joined together into a population of combined vectors;
- (d) introducing said population of combined vectors into a compatible host under conditions sufficient for expressing said population of random peptides;
- (e) determining the peptide which binds to said ligand binding protein;
- (f) isolating the nucleic acid encoding said peptide;
- and
- (g) sequencing said nucleic acid.
A method of constructing a diverse population of vectors containing expressible oligonucleotides having a desirable bias of random codon sequences, comprising operationally linking a diverse population of oligonucleotides having a desirable bias of random codon sequences to expression elements.
A method of selecting a peptide capable of being bound by a binding protein from a population of random peptides, comprising:
- (a) operationally linking a diverse population of oligonucleotides having a desirable bias of random codon sequences to expression elements;
- (b) introducing said population of vectors into a compatible host under conditions sufficient for expressing said population of random peptides;
- and
- (c) determining the peptide which binds to said ligand binding protein.
A method of determining the nucleic acid sequence encoding a peptide capable of being bound by a ligand binding protein which is selected from a population of random peptides, comprising:
- (a) operationally linking a diverse population of oligonucleotides having a desirable bias of random codon sequences to expression elements.
- (b) introducing said population of vectors into a compatible host under conditions sufficient for expressing said population of random peptides;
- (c) determining the peptide which binds to said ligand binding protein;
- (d) isolating the nucleic acid encoding said peptide;
- and
- (e) sequencing said nucleic acid.
A vector comprising two copies of a gene encoding a filamentous bacteriophage coat protein, both copies encoding substantially the same amino acid sequence but having different nucleotide sequences.
A vector comprising two copies of a gene encoding a filamentous bacteriophage coat protein, one copy of said gene capable of being operationally linked to an oligonucleotide wherein said oligonucleotide can be expressed as a fusion protein on the surface of said filamentous bacteriophage or as a soluble peptide.
Dependencies are inferred from the claim text.