Identification of 5'-upstream region of pufferfish ribosomal protein L29 gene as a strong constitutive promoter to drive GFP expression in zebrafish.
The genomic structure of Tetraodon fluviatilis L29 gene was determined and its promoter activity was analyzed in COS-1 cells and zebrafish embryos. The TfL29 gene comprises four exons and three introns, spanning approximately 1.7kb. The 5(')-upstream 2.2-kb of the first exon contains 10 E-boxes and many putative binding motifs for transcription factors GATA-1, AML-1a, c-Myb, Oct-1, CdxA, and NRF-2. Promoter activity assay showed that the distal 2.2-kb fragment not only had high luciferase activity in COS-1 cells, but also strong and ubiquitous GFP expression in a variety of tissues in zebrafish embryos. On the other hand, there are no TATA or CAAT boxes within a 300-bp region upstream from the transcription initiation site. Although this region has high luciferase activity in COS-1 cells, it is not sufficient to drive GFP expression in zebrafish embryos. In this proximal 300-bp region, there are two E-boxes, two CdxA sites, and one NRF-2 site that is immediately downstream of the transcription start site.
Chang
Ming-Huang
MH
Graduate Institute of Veterinary Medicine, National Taiwan University, Taipei, Taiwan, TOC.
Chou
Chih-Ming
CM
Hsieh
Yueh-Chun
YC
Lu
I-Ching
IC
Devi
M Kothai Nachiar
MK
Chang
Juei-Pei
JP
Kuo
Tzong-Fu
TF
Huang
Chang-Jen
CJ
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D019556:COS Cells; D017173:Caenorhabditis elegans; D004330:Drosophila; D005786:Gene Expression Regulation; D049452:Green Fluorescent Proteins; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics* / Q000378:metabolism*; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D016601:RNA-Binding Proteins; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012269:Ribosomal Proteins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D016415:Sequence Alignment; D020539:Sequence Analysis, Protein; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity; D013329:Structure-Activity Relationship; D023721:Tetraodontiformes; D015533:Transcriptional Activation; D015027:Zebrafish
D008164:Luminescent Proteins; D016601:RNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D012269:Ribosomal Proteins; C084343:Rpl29 protein, mouse; D049452:Green Fluorescent Proteins