title: Overexpression of an endo-1,4-β-glucanase V gene (EGV) from Trichoderma reesei leads to the accumulation of cellulase activity in transgenic rice.
abstract: The ectopic expression of cellulase in biomass can reduce the cost of biofuel conversion. This trait modification technique is highly beneficial for biofuel production. In this study, we isolated an endo-1,4-beta-glucanase gene (EGV) from Trichoderma reesei and inserted this gene downstream of a fragment encoding the signal peptide Apo-SP in a modified pCAMBIA1301 vector to obtain an Apo-SP and AsRed fusion protein. Transient expression of this fusion protein in onion epidermal cells showed that the Apo-SP signal was localized to the plastids. EGV transgenic rice plants that did not carry screening marker genes were obtained through overexpression of the pDTB double T-DNA vector. Western blotting showed that EGV was expressed in the dry straw of T0 generation transgenic rice plants and in fresh leaves of the T1 generation. More importantly, our results also showed that the peptide product of EGV in the transgenic plants folded correctly and was capable of digesting the cellulase substrate CMC. Additionally, cellulase activity remained stable in the straw that had been dried at room temperature for three months. This study presents an important technical approach for the development of transgenic rice straw that has stable cellulase activity and can be used for biofuel conversion.
authors.affiliation: Key Laboratory of Crop Biology of Anhui Province, Anhui Agricultural University, Hefei, China.
authors.forename: X Y
authors.initials: XY
authors.lastname: Li
authors.affiliation: Key Laboratory of Crop Biology of Anhui Province, Anhui Agricultural University, Hefei, China.
authors.forename: F
authors.initials: F
authors.lastname: Liu
authors.affiliation: Key Laboratory of Crop Biology of Anhui Province, Anhui Agricultural University, Hefei, China.
authors.forename: Y F
authors.initials: YF
authors.lastname: Hu
authors.affiliation: Beijing Weiming-Kaituo Agro-biotechnology Co., Ltd., Beijing, China.
authors.forename: M
authors.initials: M
authors.lastname: Xia
authors.affiliation: Key Laboratory of Crop Biology of Anhui Province, Anhui Agricultural University, Hefei, China.
authors.forename: B J
authors.initials: BJ
authors.lastname: Cheng
authors.affiliation: Key Laboratory of Crop Biology of Anhui Province, Anhui Agricultural University, Hefei, China.
authors.forename: S W
authors.initials: SW
authors.lastname: Zhu
authors.affiliation: Key Laboratory of Crop Biology of Anhui Province, Anhui Agricultural University, Hefei, China.
authors.forename: Q
authors.initials: Q
authors.lastname: Ma
mesh_terms: D056804:Biofuels; D002480:Cellulase / Q000096:biosynthesis* / Q000235:genetics*; D004269:DNA, Bacterial / Q000235:genetics; D018506:Gene Expression Regulation, Plant; D012275:Oryza / Q000235:genetics*; D030821:Plants, Genetically Modified / Q000235:genetics*; D014242:Trichoderma / Q000201:enzymology / Q000235:genetics
chemical_list: D056804:Biofuels; D004269:DNA, Bacterial; C036483:T-DNA; D002480:Cellulase