title: Improvement of human lysozyme expression in transgenic rice grain by combining wheat (Triticum aestivum) puroindoline b and rice (Oryza sativa) Gt1 promoters and signal peptides.
abstract: Heterologous protein expression levels in transgenic plants are of critical importance in the production of plant-made pharmaceuticals (PMPs). We studied a puroindoline b promoter and signal peptide (Tapur) driving human lysozyme expression in rice endosperm. The results demonstrated that human lysozyme expressed under the control of the Tapur cassette is seed-specific, readily extractable, active, and properly processed. Immuno-electron microscopy indicated that lysozyme expressed from this cassette is localized in protein bodies I and II in rice endosperm cells, demonstrating that this non-storage promoter and signal peptide can be used for targeting human lysozyme to rice protein bodies. We successfully employed a strategy to improve the expression of human lysozyme in transgenic rice grain by combining the Tapur cassette with our well established Gt1 expression system. The results demonstrated that when the two expression cassettes were combined, the expression level of human lysozyme increased from 5.24 +/- 0.34 mg(-1) g flour for the best single cassette line to 9.24 +/- 0.06 mg(-1) g flour in the best double cassette line, indicating an additive effect on expression of human lysozyme in rice grain.
authors.affiliation: Ventria Bioscience, 4110 North Freeway, Sacramento, CA 95834, USA.
authors.forename: Kevin
authors.initials: K
authors.lastname: Hennegan
authors.forename: Daichang
authors.initials: D
authors.lastname: Yang
authors.forename: Diane
authors.initials: D
authors.lastname: Nguyen
authors.forename: Liying
authors.initials: L
authors.lastname: Wu
authors.forename: Jeff
authors.initials: J
authors.lastname: Goding
authors.forename: Jianmin
authors.initials: J
authors.lastname: Huang
authors.forename: Fengli
authors.initials: F
authors.lastname: Guo
authors.forename: Ning
authors.initials: N
authors.lastname: Huang
authors.forename: Simon C
authors.initials: SC
authors.lastname: Watkins
mesh_terms: D001483:Base Sequence; D004274:DNA, Recombinant / Q000235:genetics; D015870:Gene Expression; D020869:Gene Expression Profiling; D005983:Glutens / Q000235:genetics; D006801:Humans; D066298:In Vitro Techniques; D009113:Muramidase / Q000096:biosynthesis* / Q000235:genetics* / Q000302:isolation & purification; D012275:Oryza / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism*; D010940:Plant Proteins / Q000235:genetics; D030821:Plants, Genetically Modified; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic; D021382:Protein Sorting Signals / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D014018:Tissue Distribution; D014908:Triticum / Q000235:genetics* / Q000378:metabolism
chemical_list: D004274:DNA, Recombinant; D010940:Plant Proteins; D021382:Protein Sorting Signals; D011994:Recombinant Proteins; C082942:puroindoline protein, Triticum aestivum; D005983:Glutens; D009113:Muramidase