Rapid isolation of promoter sequences by TAIL-PCR: the 5'-flanking regions of Pal and Pgi genes from yams (Dioscorea).
Using a modified TAIL-PCR technique, the 5'-flanking regions of the phenylalanine ammonia lyase (Pal) genes of a yam species, Dioscorea bulbifera, and the phosphoglucose isomerase (Pgi) gene of D. tokoro were successfully isolated. Two novel modifications of the TAIL-PCR procedure introduced here, namely (1) the use of a battery of random 10-mers (RAPD primers) as short arbitrary primers, and (2) the use of a total of five nested, gene-specific primers, allow the rapid isolation of the 5'-flanking region of any gene from organisms with large genomes. Isolated 5'-flanking regions were fused to the gus gene, and tested for transient expression in tobacco BY2 cells. All the isolated 5'-flanking regions were shown to drive reporter gene expression. Three Pal promoters responded to salicylic acid, presumably as a result of the binding of a MYB transcriptional activator to the multiple MREs (Myb Recognition Elements) present in these regions.
Terauchi
R
R
Plant Molecular Biology, Biocentre, University of Frankfurt, Frankfurt am Main, Germany.
Kahl
G
G
D001483:Base Sequence; D005091:Exons; D005956:Glucose-6-Phosphate Isomerase / Q000235:genetics*; D007438:Introns; D019668:Liliaceae / Q000235:genetics*; D008957:Models, Genetic; D008969:Molecular Sequence Data; D010650:Phenylalanine Ammonia-Lyase / Q000235:genetics*; D016133:Polymerase Chain Reaction / Q000379:methods*; D011401:Promoter Regions, Genetic; D017422:Sequence Analysis, DNA / Q000379:methods*; D012689:Sequence Homology, Nucleic Acid
D010650:Phenylalanine Ammonia-Lyase; D005956:Glucose-6-Phosphate Isomerase