Downregulation of Cinnamyl-Alcohol Dehydrogenase in Switchgrass by RNA Silencing Results in Enhanced Glucose Release after Cellulase Treatment

Authors

Aaron J. Saathoff, Grain, Forage, and Bioenergy Research UnitFollow
Gautam Sarath, Grain, Forage, and Bioenergy Research UnitFollow
Elaine K. Chow, Genomics and Gene Discovery Research Unit
Bruce S. Dien, USDA-ARSFollow
Christian M. Tobias, Genomics and Gene Discovery Research UnitFollow

ORCID IDs

Gautam Sarath

Document Type

Article

Date of this Version

2011

Citation

Saathoff AJ, Sarath G, Chow EK, Dien BS, Tobias CM (2011) Downregulation of Cinnamyl-Alcohol Dehydrogenase in Switchgrass by RNA Silencing Results in Enhanced Glucose Release after Cellulase Treatment. PLoS ONE 6(1): e16416. doi:10.1371/journal.pone.0016416

Abstract

Cinnamyl alcohol dehydrogenase (CAD) catalyzes the last step in monolignol biosynthesis and genetic evidence indicates CAD deficiency in grasses both decreases overall lignin, alters lignin structure and increases enzymatic recovery of sugars. To ascertain the effect of CAD downregulation in switchgrass, RNA mediated silencing of CAD was induced through Agrobacterium mediated transformation of cv. ‘‘Alamo’’ with an inverted repeat construct containing a fragment derived from the coding sequence of PviCAD2. The resulting primary transformants accumulated less CAD RNA transcript and protein than control transformants and were demonstrated to be stably transformed with between 1 and 5 copies of the TDNA. CAD activity against coniferaldehyde, and sinapaldehyde in stems of silenced lines was significantly reduced as was overall lignin and cutin. Glucose release from ground samples pretreated with ammonium hydroxide and digested with cellulases was greater than in control transformants. When stained with the lignin and cutin specific stain phloroglucinol- HCl the staining intensity of one line indicated greater incorporation of hydroxycinnamyl aldehydes in the lignin.