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DNA-based diagnostic methods have already gained acceptance in several applications.The presently important one, DNA barcoding, a species-specific marker is essential for further plant part identification as traditional medicines, such as with leaves, short trunks, pods,chopped plants or powdered plants, processed foods, fossil remains, or herbarium specimens which possess highly degraded DNA or a short reliable DNA region.The principle of DNA barcode using as a diagnostic tool, is a single piece/several pieces of DNA sequences of genes or non-coding regions to identify a life form on earth.The regions are in nucleus,mitochondrial and chloroplast genomes.The popular regions used for plants identification are in chloroplast DNA for instances rpoC1, rpoB, trnH-psbA spacer, matK and rbcL.Two options: rpoC1, matK and trnH-psbA spacer, and rpoB, matK and trnH-psbA spacer were guided to use since 2007.Two years later core barcodes include matK and rbcL were suggested for barcoding in all land plants.DNA barcodes of many plant groups in Thailand namely Aerides, Allamanda, Cinnamomum, Cissus, Cymbidium, Gardenia, Hoya, Mentha,Nymphaea, Ocimum, Piper, Pistia, Rhynchostylis, Smilax, Solanaceae, Vanda, and etc.have been constructed following these three quoted options at Molecular systematics Laboratory,Department of Biology, Faculty of Science, Khon Kaen University, with more than a thousand barcode accessions kept at GenBank.The conclusion of the most effective barcode region is trnH-psbA spacer, the second is rbcL region.Therefore, the combinations of these two regions are utmost effectiveness.For examples, in 14 Senna species, nucleotide variation of the species levels are 0.040 to 0.600 for trnH-psbA spacer and 0.010 to 0.063 of rbcL gene region, respectively.The combination values of the species level of the two regions are 0.018-0.185 indicating the utmost effectiveness.