Method of Reducing Acetylation in Plants to Improve Biofuel Production

Abstract

The invention provides methods for engineering plants to have reduced levels of acetylation by decreasing expression of one or more Cas1L genes. Such plants can be used, e.g., to increase yield for biofuel production.

What is claimed is: 
     
         1 . A method of improving the final yield from, or efficiency of, a fermentation reaction comprising plant material that is acetylated, the method comprising:
 enzymatically or chemically degrading plant material from a mutant CAS1L plant that has decreased activity of at least one CAS1L gene to obtain degradation products; and fermenting the degradation products in a fermentation reaction, wherein the final yield, or efficiency of, the fermentation reaction is increased relative to the final yield, or efficiency, of a fermentation reaction using corresponding plant material from a wildtype CAS1L plant.   
     
     
         2 . The method of  claim 1 , wherein the steps of degrading the plant material and fermenting the degradation products occur in the same reaction mixture. 
     
     
         3 . The method of  claim 1 , wherein the steps of degrading the plant material and fermenting the degradation products occur in separate reaction mixtures. 
     
     
         4 . The method of  claim 1 , wherein the improvement of the efficiency of the fermentation reaction is an increase in the amount of enzymatically degraded degradation product obtained per unit enzyme over a period of time compared to the amount of product obtained using corresponding plant material from a wildtype CAS1L plant per unit enzyme over the same period of time. 
     
     
         5 . The method of  claim 1 , wherein the mutant Cas1L plant has been engineered to decrease the activity of at least two Cas1L genes. 
     
     
         6 . The method of  claim 1 , wherein the mutant Cas1L_plan has been engineered to decrease the activity of at least three Cas1L genes. 
     
     
         7 . The method of  claim 1 , wherein the plant is a transgenic plant that comprises a vector comprising a nucleic acid sequence that encodes an RNAi that inhibits expression of at least one Cas1L gene. 
     
     
         8 . The method of  claim 1 , wherein the plant is selected from the group consisting of corn, sorghum, millet, miscanthus, sugarcane, poplar, pine, eucalyptus, wheat, rice, soy, cotton, barley, switchgrass, turfgrass, ryegrass, bahiagrass, bermudagrass, dallisgrass, pangolagrass, big bluestem, Indian grass, fescue,  Dactylis  sp.  Brachypodium,  smooth bromegrass, orchardgrass, Kentucky bluegrass, timothy, Kochia, forage soybeans, alfalfa, clover, hemp, kenaf, tobacco, potato, bamboo, rape, sugar beet, sunflower, willow, and eucalyptus. 
     
     
         9 . A plant comprising a vector that comprises an RNAi that inhibits a Cas1L gene. 
     
     
         10 . The plant of  claim 9 , wherein the plant is selected from the group consisting of corn, sorghum, millet, miscanthus, sugarcane, poplar, pine, eucalyptus, wheat, rice, soy, cotton, barley, switchgrass, turfgrass, ryegrass, bahiagrass, bermudagrass, dallisgrass, pangolagrass, big bluestem, Indian grass, fescue,  Dactylis  sp.  Brachypodium,  smooth bromegrass, orchardgrass, Kentucky bluegrass, timothy, Kochia, forage soybeans, alfalfa, clover, hemp, kenaf, tobacco, potato, bamboo, rape, sugar beet, sunflower, willow, and eucalyptus. 
     
     
         11 . An isolated mutant Cas1L plant in which expression of at least two Cas1L genes is inhibited. 
     
     
         12 . The plant of  claim 11 , wherein the plant is selected from the group consisting of corn, sorghum, millet, miscanthus, sugarcane, poplar, pine, eucalyptus, wheat, rice, soy, cotton, barley, switchgrass, turfgrass, ryegrass, bahiagrass, bermudagrass, dallisgrass, pangolagrass, big bluestem, Indian grass, fescue,  Dactylis  sp.  Brachypodium,  smooth bromegrass, orchardgrass, Kentucky bluegrass, timothy, Kochia, forage soybeans, alfalfa, clover, hemp, kenaf, tobacco, potato, bamboo, rape, sugar beet, sunflower, willow, and eucalyptus. 
     
     
         13 . A method of engineering a plant to reduce acetylation in the cell wall of a plant, the method comprising inhibiting expression of at least one Cas1L gene in the plant using RNAi. 
     
     
         14 . The method of  claim 13 , further comprising determining the level of acetylation in the cell wall of the plant. 
     
     
         15 . Bulk harvested material comprising material from at least two engineered plants that have decreased expression of at least one Cas1L gene. 
     
     
         16 . The bulk harvested material of  claim 15 , wherein the at least two engineered plants are grass plants. 
     
     
         17 . The bulk harvested material of  claim 15 , wherein the material is present in a fermentation reaction. 
     
     
         18 . A plant engineered to have reduced expression of at least one Cas1L gene, wherein the plant has at least a 10% reduction in cell wall acetylation in comparison to a plant that has not been engineered. 
     
     
         19 . The plant of  claim 18 , wherein the plant is a grass. 
     
     
         20 . A polysaccharide or polysaccharide fraction isolated from a plant of  claim 18 . 
     
     
         21 . A polysaccharide of  claim 20 , where the polysaccharide is homogalacturonan or pectin. 
     
     
         22 . A food ingredient containing the polysaccharide of  claim 20 . 
     
     
         23 . A food product containing an ingredient according to  claim 22 .