Construction Of Fish DNA Barcode Database And Excavation Of Crypticial Species In Henan Province

Located in the North China Plain of seven geographical divisions in China Henan Province has plentiful river systems.There are four major river systems,including the Yangtze River,Yellow River,Huaihe River and Haihe River,with a total number of about 1500 rivers.Most of the rivers originate from the mountains in the west,northwest,and southwest,and they present a pattern of radioactive diversion from the west to the southeast and northeast under the influence of the terrain and the trend of the mountains.According to Fish Records of Henan Province published in 1984 and Henan Fish Addendum,there are 116 species of fish in Henan province,belonging to 68 genera,16 families and 8 orders.Because of the special geographical location and changeable climatic environment,there has a variety of fish species and abundant resources.Therefore,it has an important ecological and scientific significance to have a comprehensive assessment of fish resources in Henan Province,Importantly,fish identification is the basis for the evaluation of fish resource diversity.DNA barcode technology is gradually being applied to species identification to reduce the identification errors caused by these factors.This technology uses a standardized short gene fragment with sufficient variation to quickly and accurately identify species.As a supplement and correction tool for traditional morphological identification methods,DNA barcode technology focuses on the combination of traditional morphological classification methods,and has the advantages of high efficiency,low cost,standardization and convenience.This technology can become a common technique for effectively identifying species at the “species level”.With the continuous improvement of technology and methods,DNA barcode technology is also often used to explore the discovery of species of crypticial germplasm resources and research the population genetic diversity.By constructing the DNA barcode database of Henan Province,the species diversity of fish species in Henan Province was explored,and the new species of Cobites were described by the analysis results.The main results are as follows:1.The COI genes of 89 species of fish in Henan Province were amplified and 1069 molecular data were obtained.Based on the data obtained,a DNA barcode database of fish in Henan Province was constructed.And the species coverage reached 80% of the known fish species in Henan Province.The results showed that most of the species can form single-line clusters and have high support rate,but there may have intersection speciesfor lack of phylogenetic signals or for other reasons.In the NJ phylogenetic tree,the intersection of species mainly occurs in the fishes of the Acheilognathinae and the Gobioninae.There are also a few individuals intersecting in the relationships between the species of the Triplophysa,mainly between Triplophysa pappenheimi and Triplophysa robusta.In this study,we evaluated the status of fish resources in Henan Province by analyzing the COI gene sequence,in order to provide some reference data for the later research of fish resources in Henan Province.In addition,the sequencing data obtained in this study can provide support for the construction of fish resource database in Henan Province,which is conducive to the protection of characteristic fish and the excavation of hidden germplasm resources.2.Using DNA barcode technology,we systematically analyzed and described the suspected new species of Cobitis which was found in the investigation of fish resources in Henan Province.Combining the results of DNA barcode technical data with traditional morphological analysis,it showed that the two fishes(Cobitis duplexmacula sp.nov.and Cobitis margaritatus sp.nov.)are different from the described Gambetta's pigmentations.There has a unique pigmentation pattern on the body surface of Cobitis duplexmacula sp.nov.,and the pattern of the L3 and L5 regions is zigzag and staggered.In the new species of Cobitis margaritatus sp.nov.four garge stripes were found in males,but under normal circumstances,there is no L2.L1 has ranged from 13 to 19 spacing narrower saddle brown plaques.L1 has 13 to 19 spacing narrower saddle brown plaques.In the sample,we found that the male has a black band across the longitudinal axis of the body in the L3 region,while the L3 region in the female was a black stripe at the front of the dorsal fin,and then extends backward in some irregular shape and crosses with the L1 region.Moreover,The results of the phylogenetic tree established by the two fish and their related species showed that they separately separated the one with high support rate,thus verifying the morphological identification results.