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Abstract

The invention provides a method for generating a polynucleotide sequence or population of sequences from parent polynucleotide sequences, the method comprising the steps of (a) providing a first population of polynucleotide molecules and a second population of polynucleotide molecules, the first and second populations together constituting plus and minus strands of parent polynucleotide sequences, (b) digesting the first and second populations of polynucleotide molecules with a nuclease to generate polynucleotide fragments, (c) contacting said polynucleotide fragments generated from the plus strands with fragments generated from the minus strands and (d) amplifying the fragments that anneal to each other to generate at least one polynucleotide sequence encoding one or more protein motifs having an altered amino acid sequence as compared to those encoded by the parent polynucleotides, wherein the degree of sequence variability in a selected region of the at least one polynucleotide molecule produced in step (d) is controlled by the addition of one or more oligonucleotides of predetermined variability, which oligonucleotides anneal to a sequence that lies between, but excludes, the 3′ or 5′ terminal nucleotide of the parent polynucleotide molecule. The invention also provides polynucleotides obtained by the method of the invention and polypeptides encoded by the same.

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First Claim