ECB deacylase mutants
Abstract
A method for in vitro mutagenesis and recombination of polynucleotide sequences based on polymerase-catalyzed extension of primer oligonucleotides is disclosed. The method involves priming template polynucleotide(s) with random-sequences or defined-sequence primers to generate a pool of short DNA fragments with a low level of point mutations. The DNA fragments are subjected to denaturization followed by annealing and further enzyme-catalyzed DNA polymerization. This procedure is repeated a sufficient number of times to produce full-length genes which comprise mutants of the original template polynucleotides. These genes can be further amplified by the polymerase chain reaction and cloned into a vector for expression of the encoded proteins.
- Inventors:
-
- Pasadena, CA
- Penzberg, DE
- San Diego, CA
- Sunnyvale, CA
- Issue Date:
- Research Org.:
- California Institute of Technology (CalTech), Pasadena, CA (United States)
- OSTI Identifier:
- 874307
- Patent Number(s):
- 6361988
- Assignee:
- California Institute of Technology (Pasadena, CA)
- Patent Classifications (CPCs):
-
C - CHEMISTRY C12 - BIOCHEMISTRY C12N - MICROORGANISMS OR ENZYMES
- DOE Contract Number:
- FG02-93CH10578
- Resource Type:
- Patent
- Country of Publication:
- United States
- Language:
- English
- Subject:
- ecb; deacylase; mutants; method; vitro; mutagenesis; recombination; polynucleotide; sequences; based; polymerase-catalyzed; extension; primer; oligonucleotides; disclosed; involves; priming; template; polynucleotides; random-sequences; defined-sequence; primers; generate; pool; dna; fragments; level; mutations; subjected; denaturization; followed; annealing; enzyme-catalyzed; polymerization; procedure; repeated; sufficient; times; produce; full-length; genes; original; amplified; polymerase; chain; reaction; cloned; vector; expression; encoded; proteins; dna fragment; /435/999/
Citation Formats
Arnold, Frances H, Shao, Zhixin, Zhao, Huimin, and Giver, Lorraine J. ECB deacylase mutants. United States: N. p., 2002.
Web.
Arnold, Frances H, Shao, Zhixin, Zhao, Huimin, & Giver, Lorraine J. ECB deacylase mutants. United States.
Arnold, Frances H, Shao, Zhixin, Zhao, Huimin, and Giver, Lorraine J. Tue .
"ECB deacylase mutants". United States. https://www.osti.gov/servlets/purl/874307.
@article{osti_874307,
title = {ECB deacylase mutants},
author = {Arnold, Frances H and Shao, Zhixin and Zhao, Huimin and Giver, Lorraine J},
abstractNote = {A method for in vitro mutagenesis and recombination of polynucleotide sequences based on polymerase-catalyzed extension of primer oligonucleotides is disclosed. The method involves priming template polynucleotide(s) with random-sequences or defined-sequence primers to generate a pool of short DNA fragments with a low level of point mutations. The DNA fragments are subjected to denaturization followed by annealing and further enzyme-catalyzed DNA polymerization. This procedure is repeated a sufficient number of times to produce full-length genes which comprise mutants of the original template polynucleotides. These genes can be further amplified by the polymerase chain reaction and cloned into a vector for expression of the encoded proteins.},
doi = {},
journal = {},
number = ,
volume = ,
place = {United States},
year = {Tue Jan 01 00:00:00 EST 2002},
month = {Tue Jan 01 00:00:00 EST 2002}
}
Works referenced in this record:
Synthesis of new analogs of echinocandin B by enzymatic deacylation and chemical reacylation of the echinocandin B peptide: Synthesis of the antifungal agent cilofungin (LY121019).
journal, January 1989
- Debono, M.; Abbott, B. J.; Fukuda, D. S.
- The Journal of Antibiotics, Vol. 42, Issue 3
Deacylation of echinocandin B by Actinoplanes utahensis.
journal, January 1989
- Boeck, La Verne D.; Fukuda, David S.; Abbott, Bernard J.
- The Journal of Antibiotics, Vol. 42, Issue 3