Random Mutagenesis And Amplification Of Nucleic Acid

Abstract

A method is provided for mutagenizing nucleic acids and proteins relative to an initial nucleic acid sequence by the insertion, deletion or substitution of nucleotide(s) in the target nucleic acid during amplification.

Description

CROSS-REFERENCE[0001]This application is a divisional application of application Ser. No. 10 / 069,442, filed Jun. 28, 2008, which is a 371 of PCT / US00 / 22078, filed on Aug. 11, 2000, which is a continuation-in-part of application Ser. No. 09 / 374,274, filed on Aug. 13, 1999, now U.S. Pat. No. 6,251,604, which are incorporated herein by reference in their entirety.BACKGROUND OF THE INVENTION[0002]1. Field of the Invention[0003]The present invention relates to methods for mutagenizing nucleic acids and proteins. More particularly, the present invention relates to methods for mutagenizing nucleic acids and proteins relative to an initial target nucleic acid sequence by the insertion, deletion, or substitution of nucleotide(s) in the target nucleic acid during amplification.[0004]2. Description of Related Art[0005]The sequences of genes encoding many important proteins have been determined at a rapid speed owing to the fast progress in the field of genomics. The three-dimensional structure...

AI Technical Summary

Benefits of technology

[0057]The library of oligonucleotides may optionally be synthetic and may be synthesized by randomly incorporating A, T, G, C, I or U. Optionally, at least one of the oligonucleotides used in the library of oligonucleotide in the above methods has one or more inosine residues at the 3′ end of the oligonucleotide, preferably 1-5 inosine residues, more preferably 2-4 inosine residues and most preferably 2 inosine residues. Incorporation of inosine into the oligonucleotide at the 3′ end is believed to enhance degeneracy of the oligonucleotide and promote heterologous binding of the oligonucleotide to the target sequence, which should increase the efficiency of the extension of the oligonucleotide by DNA polymerase.

Problems solved by technology

However, to a large extent, the correlation between protein primary sequence, tertiary structure, and biological function remains elusive.

Although it is desirable to be able to predict protein folding pattern from its primary sequence and to correlate its structure with function in vivo, in reality, this has proven to be a formidable task.

It is an important, but cumbersome approach to compiling an overall picture of protein functional character, let alone stability and regulatory characteristics in vivo.

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