Application of Adh Genes in the Identification of Leguminous and Gramineous Species

Recently DNA barcodes have been applied worldwide in phylogeny analysis and species identification. However, chloroplast and nuclear ribosomal sequences, which are commonly used as plant DNA barcodes, usually can not distinguish close-related species. To complement the insufficient resolution of current plant DNA barcodes, we analyzed sequence variations of Adh genes among adzuki bean vatieties, and then extended to investigate leguminous and other gramineous species, hoping to develop high-resolution Adh DNA barcode for Leguminosae and Gramineae speciess. The Adh2 gene of adzuki bean was successfuly sequenced by the FPNI-PCR method. Its gene structure is similar to those of published Adh genes, including the number and the length of exons, and the similarity of coding region (above 70%). Adh1 and Adh2 genes of nine adzuki bean varieties developed in Taiwan were also sequenced and aligned. It suggested that the Adh2 gene had higher potential to distinguish plants within species. Based on the variations of Adh DNA sequences, we developed two Adh DNA barcodes to amplify Adh1 and Adh2 genes of leguminous species and additional two Adh DNA barcodes to amplify Adh1 genes of gramineous species. These Adh DNA barcodes were successfully used to reconstruct phylogenetic tree and identify plants at inter-species and intra-species level. These demonstrated that Adh DNA barcodes complete the DNA barcode system at low taxonomic level. In addition, we discovered that paralogous and orthologous Adh genes were constantly hard to be distinguished and these cause difficulties in phylogeny analysis and species identification. Therefore, the possible causes and improving methods were also discussed in this study.