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Title: Methods for transforming and expression screening of filamentous fungal cells with a DNA library

Abstract

The present invention relates to methods for expression screening of filamentous fungal transformants, comprising: (a) isolating single colony transformants of a DNA library introduced into E. coli; (b) preparing DNA from each of the single colony E. coli transformants; (c) introducing a sample of each of the DNA preparations of step (b) into separate suspensions of protoplasts of a filamentous fungus to obtain transformants thereof, wherein each transformant contains one or more copies of an individual polynucleotide from the DNA library; (d) growing the individual filamentous fungal transformants of step (c) on selective growth medium, thereby permitting growth of the filamentous fungal transformants, while suppressing growth of untransformed filamentous fungi; and (e) measuring activity or a property of each polypeptide encoded by the individual polynucleotides. The present invention also relates to isolated polynucleotides encoding polypeptides of interest obtained by such methods, to nucleic acid constructs, expression vectors, and recombinant host cells comprising the isolated polynucleotides, and to methods of producing the polypeptides encoded by the isolated polynucleotides.

Inventors:
; ; ;
Issue Date:
Research Org.:
National Renewable Energy Lab. (NREL), Golden, CO (United States)
Sponsoring Org.:
USDOE
OSTI Identifier:
1183451
Patent Number(s):
9045748
Application Number:
11/140,150
Assignee:
Novozymes, Inc. (Davis, CA)
Patent Classifications (CPCs):
C - CHEMISTRY C40 - COMBINATORIAL TECHNOLOGY C40B - COMBINATORIAL CHEMISTRY
C - CHEMISTRY C12 - BIOCHEMISTRY C12N - MICROORGANISMS OR ENZYMES
DOE Contract Number:  
AC36-98GO10337
Resource Type:
Patent
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; 60 APPLIED LIFE SCIENCES

Citation Formats

Teter, Sarah, Lamsa, Michael, Cherry, Joel, and Ward, Connie. Methods for transforming and expression screening of filamentous fungal cells with a DNA library. United States: N. p., 2015. Web.
Teter, Sarah, Lamsa, Michael, Cherry, Joel, & Ward, Connie. Methods for transforming and expression screening of filamentous fungal cells with a DNA library. United States.
Teter, Sarah, Lamsa, Michael, Cherry, Joel, and Ward, Connie. Tue . "Methods for transforming and expression screening of filamentous fungal cells with a DNA library". United States. https://www.osti.gov/servlets/purl/1183451.
@article{osti_1183451,
title = {Methods for transforming and expression screening of filamentous fungal cells with a DNA library},
author = {Teter, Sarah and Lamsa, Michael and Cherry, Joel and Ward, Connie},
abstractNote = {The present invention relates to methods for expression screening of filamentous fungal transformants, comprising: (a) isolating single colony transformants of a DNA library introduced into E. coli; (b) preparing DNA from each of the single colony E. coli transformants; (c) introducing a sample of each of the DNA preparations of step (b) into separate suspensions of protoplasts of a filamentous fungus to obtain transformants thereof, wherein each transformant contains one or more copies of an individual polynucleotide from the DNA library; (d) growing the individual filamentous fungal transformants of step (c) on selective growth medium, thereby permitting growth of the filamentous fungal transformants, while suppressing growth of untransformed filamentous fungi; and (e) measuring activity or a property of each polypeptide encoded by the individual polynucleotides. The present invention also relates to isolated polynucleotides encoding polypeptides of interest obtained by such methods, to nucleic acid constructs, expression vectors, and recombinant host cells comprising the isolated polynucleotides, and to methods of producing the polypeptides encoded by the isolated polynucleotides.},
doi = {},
journal = {},
number = ,
volume = ,
place = {United States},
year = {2015},
month = {6}
}

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Works referenced in this record:

Mutation and screening to increase chymosin yield in a genetically-engineered strain ofAspergillus awamori
journal, June 1990


A System for Dual Protein Expression in Pichia pastoris and Escherichia coli
journal, December 2000


High-throughput screening for soluble recombinant expressed kinases in Escherichia coli and insect cells
journal, July 2004