title: Enhanced transgene expression in rice following selection controlled by weak promoters.
abstract: BACKGROUND
  Techniques that enable high levels of transgene expression in plants are attractive for the commercial production of plant-made recombinant pharmaceutical proteins or other gene transfer related strategies. The conventional way to increase the yield of desired transgenic products is to use strong promoters to control the expression of the transgene. Although many such promoters have been identified and characterized, the increase obtainable from a single promoter is ultimately limited to a certain extent.
  
  
  RESULTS
  In this study, we report a method to magnify the effect of a single promoter by using a weak promoter-based selection system in transgenic rice. tCUP1, a fragment derived from the tobacco cryptic promoter (tCUP), was tested for its activity in rice by fusion to both a β-glucuronidase (GUS) reporter and a hygromycin phosphotransferase (HPT) selectable marker. The tCUP1 promoter allowed the recovery of transformed rice plants and conferred tissue specific expression of the GUS reporter, but was much weaker than the CaMV 35S promoter in driving a selectable marker for growth of resistant calli. However, in the resistant calli and regenerated transgenic plants selected by the use of tCUP1, the constitutive expression of green fluorescent protein (GFP) was dramatically increased as a result of the additive effect of multiple T-DNA insertions. The correlation between attenuated selection by a weak promoter and elevation of copy number and foreign gene expression was confirmed by using another relatively weak promoter from nopaline synthase (Nos).
  
  
  CONCLUSIONS
  The use of weak promoter derived selectable markers leads to a high T-DNA copy number and then greatly increases the expression of the foreign gene. The method described here provides an effective approach to robustly enhance the expression of heterogenous transgenes through copy number manipulation in rice.
authors.affiliation: State Key Laboratory Breeding Base for Zhejiang Sustainable Pest and Disease Control, MOA Key Laboratory for Plant Protection and Biotechnology, Zhejiang Provincial Key Laboratory of Plant Virology, Zhejiang Academy of Agricultural Sciences, Hangzhou, 310021, P,R,China.
authors.forename: Jie
authors.initials: J
authors.lastname: Zhou
authors.forename: Yong
authors.initials: Y
authors.lastname: Yang
authors.forename: Xuming
authors.initials: X
authors.lastname: Wang
authors.forename: Feibo
authors.initials: F
authors.lastname: Yu
authors.forename: Chulang
authors.initials: C
authors.lastname: Yu
authors.forename: Juan
authors.initials: J
authors.lastname: Chen
authors.forename: Ye
authors.initials: Y
authors.lastname: Cheng
authors.forename: Chenqi
authors.initials: C
authors.lastname: Yan
authors.forename: Jianping
authors.initials: J
authors.lastname: Chen
mesh_terms: D000594:Amino Acid Oxidoreductases / Q000235:genetics; D001483:Base Sequence; D017796:Caulimovirus / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism; D018628:Gene Dosage; D018506:Gene Expression Regulation, Plant; D005822:Genetic Vectors / Q000235:genetics / Q000378:metabolism; D005966:Glucuronidase / Q000235:genetics / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D012275:Oryza / Q000378:metabolism*; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000235:genetics / Q000378:metabolism; D010940:Plant Proteins / Q000235:genetics; D030821:Plants, Genetically Modified / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics; D014026:Nicotiana / Q000378:metabolism; D019076:Transgenes / Q000235:genetics*
chemical_list: D004269:DNA, Bacterial; D010940:Plant Proteins; D011993:Recombinant Fusion Proteins; C036483:T-DNA; D049452:Green Fluorescent Proteins; D000594:Amino Acid Oxidoreductases; C021883:nopaline synthase; D017853:Phosphotransferases (Alcohol Group Acceptor); C063242:hygromycin-B kinase; D005966:Glucuronidase