Abstract:
The specific primers of open reading frame (ORF) of dihydroflavonol(
DFR) gene in brier grapes (
Vitis davidii Foёx.) were designed according to the CDS template of the gene.
DFR gene sequence was cloned using RT-PCR, andsubsequently, the genetic characteristics analyzed by bioinformatics. The 1 014 bp full-length cDNA of
DRF's ORF was thus obtained. It encoded 337 amino acids, and was named
V. davidii DFR 4-reductase gene (
VdDFR) with GenBank accession number of KF915803. The predicted molecular weight of VdDFR protein was 37 593.2 Da,theoretical pI is 5.81. As a transmembrane and a hydrophilic protein, it did not belong to secretory category,had no signal peptide, and was located largely in the cytoplasm (70%). The secondary structure ofthe mixed protein was mostly random coil (52.82%). The amino acids sequence of the gene possibly contained 7 glycosylation sites, 16 phosphorylation sites, one NAD(P) binding site, and one NAD-dependent epimerase/dehydratase(N-terminal) domain, and the gene likely belonged to the NADB_Rossmann superfamily. The nucleotide sequences of
DFR from
V.davidii,V.bellula,V.amurensis, and
V.vinifera were 99% homogenous; those of
V.davidii and
V.rotundifolia, 98% homogenous; and, those of
V.davidii and
Ampelopsis grossedentata, 94% homogenous. These results indicated that the
DFR gene coding region was evolutionally conservative. And, the phylogenetic tree constructed based on the sequence salso reflected the same evolutionary trait of these plants.