Application Of DNA Barcoding In Umbilicaria Species

DNA Barcoding is a molecular taxonomic method based on a short standard DNA sequence, which has been developed and applied in the identification of biological specimens in the whole world. As an assistant method for the traditional taxonomy, the digital DNA barcoding has a huge significance on the species automated identification, the monitoring of biological diversity and the protection of endangered species. The Umbilicaria species are a kind of foliose lichenized fungi with big, independent thallus. Because of their wide distribution and edible and medicinal properties, Umbiliaria species have always been one of the hot research spots in the field of taxonomy.We first amplified and sequenced the ITS sequences of 305 Umbilicaria specimens by using the general primer pair ITS5 and ITS4, and obtained 228 valid ITS sequences with 74.8% of success rate. These 228 ITS sequences were used to make an alignment and produced a DNA Barcode sequence with 520 bp of valid length. On the basis of the above research, we reconstructed a taxon identification tree, and divided the 15 species into 8 big clustering clusters according to ITS sequence. We also confirmed the positions of the suspected species of U. yunnana and the new subspecies of U. hypococcinea on the taxon identification tree by using ITS sequence. Our results showed that ITS sequence is a good DNA barcode candidate for Umbilicaria species, which can not only distinguish the species of Umbilicaria easily, but also provide new clue for determination of position of suspected species on the taxon identification tree.At the same time, we also identified 24 microcode from ITS1 and ITS2 regions in 18 species of Umbilicaria, and found that the variations of only 3 or 4 bases within these microcode sequences are enough to distinguish these species. These identified microcode sequences may be used to construct species identification chips. Our results showed that microcoding can recover the imperfection of phylogenetic tree, and be useful for identification of the geographical vicariad and sister species.In conclusion, our studies on DNA barcoding will contribute to the combination of phenotypic and genotypic research of Umbilicaria.