Deletion analysis and localization of SbPRP1, a soybean cell wall protein gene, in roots of transgenic tobacco and cowpea.
SbPRP1 is a member of the soybean (Glycine max L. Merr) proline-rich cell wall protein family and is expressed at high levels in root tissue. To characterize the sequences required for this expression, we have fused 1.1 kb of upstream flanking DNA sequence from an SbPRP1 genomic clone to a gene encoding beta-glucuronidase (GUS). This construct was introduced into tobacco using Agrobacterium tumefaciens-mediated transformation. Histochemical staining of GUS activity in transgenic tobacco indicated that SbPRP1 is expressed in the apical and elongating region of both primary and lateral roots, most strongly in the epidermis. A similar localization pattern was found in transformed hairy roots when this construct was introduced into cowpea (Vigna aconitifolia) using Agrobacterium rhizogenes-mediated transformation. Nested 5'-deletion analysis of the SbPRP1 promoter indicated that a minimal promoter for SbPRP1 expression in roots is located within the first 262 bases of upstream flanking DNA and that the region between -1080 and -262 is required for maximal expression of this gene. Gel retardation assays showed that nuclear factors can be detected in soybean roots which specifically bind to sequences located between -1080 and -623, a region which is needed for maximal expression of the SbPRP1 promoter. Northern hybridization analysis was also used to show that little SbPRP1 mRNA was present in roots during the first 24 h after imbibition. These studies indicate that SbPRP1 expression is localized to the actively growing region of the root and that this expression is temporally regulated during very early stages of seedling growth.
Suzuki
H
H
Agronomy Department, Ohio Agricultural Research and Development Center, Columbus.
Fowler
T J
TJ
Tierney
M L
ML
D001483:Base Sequence; D015152:Blotting, Northern; D002473:Cell Wall / Q000378:metabolism; D007887:Fabaceae / Q000235:genetics*; D017353:Gene Deletion; D005786:Gene Expression Regulation; D005966:Glucuronidase / Q000235:genetics / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics*; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D010940:Plant Proteins / Q000235:genetics*; D030821:Plants, Genetically Modified; D010946:Plants, Medicinal; D010947:Plants, Toxic; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D015183:Restriction Mapping; D017384:Sequence Deletion; D013025:Glycine max / Q000235:genetics*; D014026:Nicotiana / Q000235:genetics*
D008565:Membrane Proteins; D009838:Oligodeoxyribonucleotides; D010940:Plant Proteins; C063058:proline-rich cell wall proteins, soybean; D012313:RNA; D005966:Glucuronidase