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Gene targeting in a nonhomologous end joining deficient Hypocrea jecorina

Paper ID Volume ID Publish Year Pages File Format Full-Text
24884 43543 2009 6 PDF Available
Title
Gene targeting in a nonhomologous end joining deficient Hypocrea jecorina
Abstract

The industrially applied ascomycete Hypocrea jecorina (synonym: Trichoderma reesei) exhibits a low rate of exogenous DNA integration by homologous recombination (HR). This hinders the high-throughput generation of strains by gene replacement and is therefore impeding systematic functional gene analyses towards, e.g. strain improvement for protein or enzyme production. To increase the rate of HR events during fungal transformation we identified and deleted the orthologue of the human KU70 in H. jecorina, which is required for the nonhomologous end joining (NHEJ) pathway and responsible for ectopic DNA integration. The effect of the absence of the H. jecorina tku70 on gene targeting was tested by deletion of two so far uncharacterized genes encoding a short chain dehydrogenase and a fungal specific transcription factor. Efficiency of gene targeting for both genes was >95% in a Δtku70 strain when 1 kb homologous flanking regions were used in the deletion construct. This is a significant increase in targeting efficiency compared to the parental – non-tku70 deleted – strain TU-6 where a gene knock-out frequency of only 5–10% was observed. Together with the recently annotated genomic sequence of H. jecorina, this system provides a useful tool for a genome-wide functional gene analysis on a high-throughput scale to improve the biotechnological potential of this fungus.

Keywords
Trichoderma reesei; ku70; Gene targeting; Gene knock-out; Nonhomologous end joining
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Gene targeting in a nonhomologous end joining deficient Hypocrea jecorina
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 139, Issue 2, 15 January 2009, Pages 146–151
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us