title: Characterization of a T-DNA promoter trap line of Arabidopsis thaliana uncovers a cryptic bi-directional promoter.
abstract: Investigation of the transgenic Arabidopsis promoter trap line GFP-868 that showed GFP expression only in anthers revealed the T-DNA insertion at 461bp upstream to the hypothetical gene At4g10596 with the GFP reporter gene in head-to-head orientation to the At4g10596 gene. The expression of the At4g10596 gene in wild type and in GFP-868 plant homozygous for T-DNA insertion was comparable and found in all tissues tested, while the GFP expression was restricted to anthers of the GFP-868 plants suggesting that the 461bp fragment separating the two genes in the GFP-868 line is functioning as bi-directional promoter. This 461bp fragment was cloned upstream to the GUS gene in two orientations to test for bi-directional promoter activity. Transgenic Arabidopsis plants carrying either of these constructs showed GUS activity in anthers indicating that this fragment behaves as bi-directional promoter specific to anthers. These results were also supported by the presence of cis-acting motifs such as TATA box and POLLEN1LELAT52 (AGAAA) within the 461bp sequence in both orientations. However, transcripts corresponding to the upstream sequences beyond -461 nucleotides were not detected in the wild type suggesting that this 461bp fragment is a cryptic promoter. The significance of the promoter trap approach and the usefulness of this type of promoter are discussed.
authors.affiliation: Biotechnology Division, CSIR-Institute of Himalayan Bioresource Technology, Palampur 176061, Himachal Pradesh, India.
authors.forename: Pritu
authors.initials: P
authors.lastname: Pratibha
authors.forename: Sunil Kumar
authors.initials: SK
authors.lastname: Singh
authors.forename: Isha
authors.initials: I
authors.lastname: Sharma
authors.forename: Ravi
authors.initials: R
authors.lastname: Kumar
authors.forename: Ramamurthy
authors.initials: R
authors.lastname: Srinivasan
authors.forename: Shripad Ramachandra
authors.initials: SR
authors.lastname: Bhat
authors.forename: Paramvir Singh
authors.initials: PS
authors.lastname: Ahuja
authors.forename: Yelam
authors.initials: Y
authors.lastname: Sreenivasulu
mesh_terms: D016960:Agrobacterium tumefaciens / Q000235:genetics / Q000378:metabolism; D017360:Arabidopsis / Q000235:genetics*; D001483:Base Sequence; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000235:genetics*; D035264:Flowers / Q000235:genetics; D018506:Gene Expression Regulation, Plant; D017343:Genes, Plant; D017930:Genes, Reporter; D005966:Glucuronidase / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006720:Homozygote; D008969:Molecular Sequence Data; D030821:Plants, Genetically Modified / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic; D012045:Regulatory Sequences, Nucleic Acid; D016385:TATA Box
chemical_list: D004269:DNA, Bacterial; C036483:T-DNA; D049452:Green Fluorescent Proteins; D005966:Glucuronidase