title: Isolation of the maize Zpu1 gene promoter and its functional analysis in transgenic tobacco plants.
abstract: By screening a genomic library of maize, a 2.2 kb 5' flanking fragment of Zpu1 gene, encoding the pullulanase-type starch debranching enzyme, was isolated. Promoter fragments of various lengths, including the full 5' flanking sequence (-2267 to -1) (Z1), a 3' deletion (-2267 to -513) (Z5) and three 5' deletions extending to -1943 (Z2), -1143 (Z3) and -516 (Z4) upstream of the translational initiation codon (ATG), were fused to the GUS reporter gene and introduced into tobacco. When these constructs were tested in transgenic tobacco plants, seed-preferred GUS activity was observed in pZ1-transgenic lines. In pZ2-transgenic lines, the GUS activity was not only restricted to seeds, but was also detected in calyxes, petals, stamens and mature leaves. At the same time, negligible GUS activity was detected in roots, stems, young leaves, stigmas and ovaries from the transgenic tobacco plants, which had integrated the full isolated sequence of Zpu1 promoter or its deletions. Deletion analysis indicated that the promoter contained a putative positive cis-regulatory element and the proximal region (-516 to -1) was essential for directing the expression of gus reporter gene. Analysis of GUS activity during the fruit development and seed germination suggested that Zpu1 promoter is active both in starch anabolism and in starch catabolism, which is consistent with the function of the endogenous gene in maize. GUS activity in leaves under light and darkness confirmed that Zpu1 promoter functions in the starch degradation of photosynthetic tissues in the dark phase of the diurnal cycle.
authors.affiliation: State Key Laboratory of Agrobiotechnology and Department of Seed Sciences and Technology, China Agricultural University, Beijing 100094, China.
authors.forename: Xiaoping
authors.initials: X
authors.lastname: Chen
authors.forename: Zhangying
authors.initials: Z
authors.lastname: Wang
authors.forename: Riliang
authors.initials: R
authors.lastname: Gu
authors.forename: Junjie
authors.initials: J
authors.lastname: Fu
authors.forename: Jianhua
authors.initials: J
authors.lastname: Wang
authors.forename: Ying
authors.initials: Y
authors.lastname: Zhang
authors.forename: Maoyan
authors.initials: M
authors.lastname: Wang
authors.forename: Jinpeng
authors.initials: J
authors.lastname: Zhang
authors.forename: Jinping
authors.initials: J
authors.lastname: Jia
authors.forename: Guoying
authors.initials: G
authors.lastname: Wang
mesh_terms: D024506:5' Flanking Region / Q000235:genetics; D020029:Base Pairing; D001483:Base Sequence; D004587:Electrophoresis, Agar Gel; D018506:Gene Expression Regulation, Plant; D017343:Genes, Plant; D018525:Germination; D005966:Glucuronidase / Q000378:metabolism; D006026:Glycoside Hydrolases / Q000235:genetics*; D008969:Molecular Sequence Data; D010788:Photosynthesis; D018515:Plant Leaves / Q000235:genetics; D010940:Plant Proteins / Q000235:genetics*; D030821:Plants, Genetically Modified; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012639:Seeds / Q000502:physiology; D017422:Sequence Analysis, DNA; D014026:Nicotiana / Q000235:genetics*; D003313:Zea mays / Q000235:genetics*
chemical_list: D010940:Plant Proteins; D006026:Glycoside Hydrolases; C118576:ZPU1 protein, Zea mays; D005966:Glucuronidase