DNA barcode technology is a technique for rapid and accurate identification of species by DNA sequences. This technology provides a theoretical basis for studying the evolutionary laws, genetic variation, phylogeny and biodiversity of species. Because this technology has the use of some genetically conserved DNA fragments in biological populations for species identification and phylogenetic positioning, understanding its branch sources, and even predicting its evolutionary direction, it has become a taxonomist in recent years. Hot spots of concern.
1, the workflow
The molecular biology techniques used in DNA barcode technology are not complicated. The main workflows include sample collection, DNA extraction, design and synthesis of universal primers, primer selection, optimization of reaction conditions for PCR amplification, purification of PCR products, sequencing and analysis. . Briefly, by PCR amplification and sequencing of a set of short homologous DNA sequences (about 800 bp) from different organisms, followed by multiple sequence alignment and cluster analysis of the measured sequences, A body is precisely positioned in a taxonomic group.
Sequence data analysis is the most important part of DNA barcode exploration. Firstly, sequence alignment and manual correction are performed. The K2P distance between species and species is calculated by MEGA or PAUP, which is used to represent the degree of sequence variation between different classification orders. Sequence differences at three levels of species, genus and family, and then based on the calculation results to establish a NJ tree (neighbour-joining tree), and finally the unknown specimens can be classified and identified according to the genetic distance of the DNA barcode.
2 technical characteristics
(1) Not affected by the developmental stage. The DNA sequence information of the same organism is the same in different life cycles, so the technology detection object can be every period in the biological life process.
(2) Not affected by individual morphological characteristics. For angiosperms, it is difficult to identify correctly based on specimens lacking flowers and fruits. Classification using DNA barcode technology does not affect the recognition results even when the sample is damaged, and it can accurately identify species with high morphological similarity.
(3) Not subject to species restrictions. Once established, a standard DNA barcode database can make identification of various species possible, not limited to endangered species, indigenous species or invasive species.
(4) It is possible to identify hidden taxon units that are ubiquitous in many groups.
(5) A large amount of information is obtained. A large number of samples can be quickly identified at a time by establishing a database of DNA barcodes.
(6) High accuracy. The DNA barcode technology digitizes species identification using sequences consisting of the base ACTG. This technology has more accurate, reliable and objective characteristics than the characterization method.
(7) The operation method is simple and efficient and easy to grasp. It does not require highly accurate bio-professional techniques, making it easy for interdisciplinary researchers to use the technology to speed up the process of taxonomy.
3. Application of DNA barcode
(1) Enlargement of the scope of identification of species
Currently, DNA barcodes have been widely used for the identification and classification of various species. Chen Nian et al also studied the application of DNA barcode technology in fungal classification. Researchers in the classification and identification of animals also confirmed the feasibility and effectiveness of DNA barcode technology. Pan Chengying et al. studied the mitochondrial COI gene of seven species of mites in Catantopidae as a DNA barcode to identify aphid species. Feasibility, the results showed that the DNA classification and morphological classification of 7 species of 3 genera were basically the same.
(2) Discovery of hidden taxa
The number of species is difficult to determine, and one of the reasons is that we are not sure how many hidden species there are in nature. Hidden species are not new species. They are different species that are not classified in the traditional taxonomy and are classified as the same species. The emergence of DNA barcode technology has become an effective way to discover those species that are similar in morphology but have genetic differentiation. New species such as giant algae, Madagascar ants, and Australian fish are found in the identification of species using DNA barcode technology.
(3) Discussion on phylogenetic relationship
Sub-system development analysis refers to the study of the evolutionary relationship between species at the molecular level. It directly uses the information extracted from nucleic acid sequences or protein molecules as a feature of the species, and builds a phylogenetic tree by comparing the relationships between the sequences of biological molecules. Then clarify the evolutionary relationship between the various species. When building a barcode database, you can refer to the barcode of life data system (BOLD, http://), which has more than 460,000 records, covering more than 46,000 animals. Species, and the database is expanding.
4. Outlook
DNA barcode technology has proven to be an effective bioassay not only as a powerful complement to traditional species identification, but also because it uses digital forms to automate and standardize the sample identification process. Diversity protection provides comprehensive, accurate, fast and convenient species information services, breaking through the excessive dependence of traditional identification methods on experience. There is reason to believe that DNA barcode technology, combined with other taxonomic methods, can help identify species and speed up the discovery of new species.
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