Transient expression assays with the proximal promoter of a newly characterized actin gene from the oyster Crassostrea gigas.
We undertook the characterization of an actin gene and its proximal promoter in the oyster Crassostrea gigas. A complete actin cDNA was identified, sequenced and its amino acid sequence deduced. Comparative analysis showed a high homology with actin of other species and that this gene is closer to the cytoplasmic form of actins than to the muscle type. A probe derived from the 5'-untranslated region of the cDNA was then used to isolate the actin gene from a genomic library. The gene was sequenced and shown to contain a single 643 bp intron. A 1670 bp fragment upstream from the open reading frame was isolated and sequenced. This upstream region displays typical features of actins such as a serum response element (CarG box). This fragment was cloned into the promoterless vector pGL3-basic and the resulting construct was transfected into cells of dissociated oyster heart primary cultures. Its capacity to express the luciferase in this in vitro homologous system was monitored and showed high expression levels. This is the first complete actin sequence reported so far for the oyster C. gigas and its promoter is the first available among bivalves.
Cadoret
J P
JP
Défense et Résistance chez les Invertébrés Marins, IFREMER-CNRS-Université de Montpellier II, France. jean.paul.cadoret@ifremer.fr
Debón
R
R
Cornudella
L
L
Lardans
V
V
Morvan
A
A
Roch
P
P
Boulo
V
V
D000199:Actins / Q000737:chemistry / Q000235:genetics*; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D003001:Cloning, Molecular; D004268:DNA-Binding Proteins / Q000235:genetics; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D008969:Molecular Sequence Data; D009154:Mutation; D009687:Nuclear Proteins / Q000235:genetics; D010125:Ostreidae / Q000235:genetics*; D011401:Promoter Regions, Genetic; D026362:Serum Response Factor; D014162:Transfection
D000199:Actins; D004268:DNA-Binding Proteins; C400704:GiaI protein, Crassostrea gigas; D009687:Nuclear Proteins; D026362:Serum Response Factor