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Expression of Recombinant Cellulase Cel5A from Trichoderma reesei in Tobacco Plants

 
: Garvey, M.; Klinger, J.; Klose, H.; Fischer, R.; Commandeur, U.

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Journal of visualized experiments : JoVE. Online resource (2014), No.88, Art. e51711
http://www.jove.com/
ISSN: 1940-087X
English
Journal Article, Non Book Material
Fraunhofer IME ()

Abstract
Cellulose degrading enzymes, cellulases, are targets of both research and industrial interests. The preponderance of these enzymes in difficult-to-culture organisms, such as hyphae-building fungi and anaerobic bacteria, has hastened the use of recombinant technologies in this field. Plant expression methods are a desirable system for large-scale production of enzymes and other industrially useful proteins. Herein, methods for the transient expression of a fungal endoglucanase, Trichoderma reesei Cel5A, in Nicotiana tabacum are demonstrated. Successful protein expression is shown, monitored by fluorescence using an mCherry-enzyme fusion protein. Additionally, a set of basic tests are used to examine the activity of transiently expressed T. reesei Cel5A, including SDS-PAGE, Western blotting, zymography, as well as fluorescence and dye-based substrate degradation assays. The system described here can be used to produce an active cellulase in a short time period, so as to assess the potential for further production in plants through constitutive or inducible expression systems.

: http://publica.fraunhofer.de/documents/N-360093.html