title: Production of a Highly Protease-Resistant Fungal α-Galactosidase in Transgenic Maize Seeds for Simplified Feed Processing.
abstract: Raffinose-family oligosaccharide (RFO) in soybeans is one of the major anti-nutritional factors for poultry and livestocks. α-Galactosidase is commonly supplemented into the animal feed to hydrolyze α-1,6-galactosidic bonds on the RFOs. To simplify the feed processing, a protease-resistant α-galactosidase encoding gene from Gibberella sp. strain F75, aga-F75, was modified by codon optimization and heterologously expressed in the embryos of transgentic maize driven by the embryo-specific promoter ZM-leg1A. The progenies were produced by backcrossing with the commercial inbred variety Zheng58. PCR, southern blot and western blot analysis confirmed the stable integration and tissue specific expression of the modified gene, aga-F75m, in seeds over four generations. The expression level of Aga-F75M reached up to 10,000 units per kilogram of maize seeds. In comparison with its counterpart produced in Pichia pastoris strain GS115, maize seed-derived Aga-F75M showed a lower temperature optimum (50 °C) and lower stability over alkaline pH range, but better thermal stability at 60 °C to 70 °C and resistance to feed pelleting inactivation (80 °C). This is the first report of producing α-galactosidase in transgenic plant. The study offers an effective and economic approach for direct utilization of α-galactosidase-producing maize without any purification or supplementation procedures in the feed processing.
authors.affiliation: Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing, P. R. China.
authors.forename: Wenxia
authors.initials: W
authors.lastname: Yang
authors.affiliation: Biotechnology Institute, Chinese Academy of Agricultural Sciences, Beijing, P. R. China.
authors.forename: Yuhong
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Biotechnology Institute, Chinese Academy of Agricultural Sciences, Beijing, P. R. China.
authors.forename: Xiaojin
authors.initials: X
authors.lastname: Zhou
authors.affiliation: Biotechnology Institute, Chinese Academy of Agricultural Sciences, Beijing, P. R. China.
authors.forename: Wei
authors.initials: W
authors.lastname: Zhang
authors.affiliation: Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing, P. R. China.
authors.forename: Xiaolu
authors.initials: X
authors.lastname: Xu
authors.affiliation: Biotechnology Institute, Chinese Academy of Agricultural Sciences, Beijing, P. R. China.
authors.forename: Rumei
authors.initials: R
authors.lastname: Chen
authors.affiliation: Institute of Food Crops, Jiangsu Academy of Agricultural Sciences, Nanjing, P. R. China.
authors.forename: Qingchang
authors.initials: Q
authors.lastname: Meng
authors.affiliation: Institute of Food Crops, Jiangsu Academy of Agricultural Sciences, Nanjing, P. R. China.
authors.forename: Jianhua
authors.initials: J
authors.lastname: Yuan
authors.affiliation: Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing, P. R. China.
authors.forename: Peilong
authors.initials: P
authors.lastname: Yang
authors.affiliation: Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing, P. R. China.
authors.forename: Bin
authors.initials: B
authors.lastname: Yao
mesh_terms: D000595:Amino Acid Sequence; D000821:Animal Feed; D015139:Blotting, Southern; D015153:Blotting, Western; D003062:Codon / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D018506:Gene Expression Regulation, Plant; D005822:Genetic Vectors / Q000378:metabolism; D005874:Gibberella / Q000201:enzymology*; D008969:Molecular Sequence Data; D010447:Peptide Hydrolases / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry; D010641:Phenotype; D010843:Pichia / Q000235:genetics; D030821:Plants, Genetically Modified; D016133:Polymerase Chain Reaction; D012038:Regeneration; D012639:Seeds / Q000235:genetics*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013696:Temperature; D014170:Transformation, Genetic; D003313:Zea mays / Q000235:genetics*; D000519:alpha-Galactosidase / Q000096:biosynthesis*
chemical_list: D003062:Codon; D010455:Peptides; D000519:alpha-Galactosidase; D010447:Peptide Hydrolases