Medicinal Coptis Molecular Systematics And Protection Of Genetic Research

Coptis in China, group of three all splited palmate leaves, is divided into four species and one variant by morphology according to Flora of China, they are Coptis omeiensis(Chen) C.Y.Cheng, C.deltoidea C.Y.Cheng et Hsiao, C.teeta Wall, C.chinensis Franch and C.chinensis Franch var brevisepala W T Wanget Hsiao. The five species present dispute and confusion in taxonomy, especially in the origin of cultivation. C.chinensis and C.deltoidea are all cultivated products, with the source of wild germplasm unclear; Coptis omeiensis with the distribution in the core area of Mount. Emei, were seriously endangered; C.teeta distributed in Yunnan, Tibet high-altitude regions, is being endangered. C.chinensis Franch var brevisepala, with wider distribution area of wild germplasm, also became the endangered species with reduced amount of resources because of local substitutes for Huang Lian. Samples collected in this study included the five species, C.omeiensis and C.chinensis Franch var brevisepala are wild. C. teeta was transplanted, while Coptis chinensis and C.deltoidea were cultivated products.The present study restructed the molecular systematics with 226 samples collected, using chloroplast DNA sequence methods (cpSNP), using the two fragments of non-coding regions of chloroplast psbA-trnH and trnL-trnF, two fragments of the variable sites were 23 and 9, with 17 haplotype in all. Sequences of all samples of different populations and a evolutionary tree was built with the following conclusions:1) Coptis of group three all splited palmate leaves constructed into three clear branches, C.teeta, a branch with the farthest genetic distance; a branch contained C.omeiensis accompanied cultural products; the other branch, which keeping more haplotype, are all cultivated products.2) With the complicated geographical structure of Mount. Emei, C. omeiensis were divided into two distinct sub-branch. From the haplotype distribution in different species, there presented confusing problem of cultivated germplasm.C. deltoidea, with the highest haplotype frequency of haplotype H, are only one base variation between haplotype A of C.omeiensis and C.deltoidea in population DCZX are all haplotype A, speculated cultivation of C.deltoidea originated in C.omeiensis; which, C.deltoidea population DCZX, also called "Caolian", the lobes long similar with C.omeiensis and the same haplotype A, speculated cultivars of C.omeiensis. The long cultivated history and extensive cultivation with retained abundant haplotype, C.chinensis were in the other branch outside C.omeiensis, therefore speculated that C.chinensis cultivated from another species (the ancestor have disappeared), which domesticated by introduction and cultivated by ancients. C.chinensis var brevisepala, the haplotype with only 4-step variation between haplotype F, as sub-species of C. chinensis were confirmed by genetic variants.For this, the author dealed with ancient herbal literatur of Coptis, found that C. teeta was rarely recorded, the ancient records of Coptis began from Han and Jin, there are two kinds, which related the two cultivated species now, C.chinensis and C.deltoidea. Cultivation history recorded of Coptis, "chicken feet", that is C.chinensis, cultivated germplasm from Wuxi, Chongqing, with seed multiplication, Wushan, its complex geographic structure, suggesting that Wushan area most likely an ancient source of C.chinensis cultivated germplasm. C.deltoidea, from Hongya historical cultivation records and records of C.omeiensis distributed in Hongya, confirmed C.deltoidea cultivation originated from C.omeiensis. And many of the ancient herbal Coptis recorded distribution points were gone, accorded with the trend of wild germplasm Coptis further narrowing now.This is the first SSR primers development of medicinal plant C.omeiensis. Using five pairs of fluorescent SSR primers, the genetic diversity among populations, genetic structure and gene flow in six wild populations and one transplanted populations of C.omeiensis were studied The results show that the existence of C.omeiensis with abundant genetic diversity{A= 4.7; HO= 0.397; HE=0.588), may be related to the complex geographical structure. STRUCTURE software analysis represented two genetic C.omeiensis groups, consistent with the chloroplast analysis data. Comparison and analysis of the two data, there has been extensive gene flow between the two groups, but mainly for pollen flow, not seed flow. Using software BOTTLENECK analysis, four out of six wild populations showed over heterozygosity, these populations suffered from recent genetic bottleneck, which probable man-made excavation damage in 90's. The genetic form of C.omeiensis affected greatly after the fragmented disdribution, mainly with reduction of gene flow among populations, genetic differentiation and inbreeding depression phenomenon. Gene flow among populations decreased (Nm= 0.8812), AMOVA analysis showed that the fragments with significant genetic differentiation. In the four populations, CDC, SDH, BY and RTZ, inbreeding coefficient within populations FIS were high and significantly deviated from Hardy-Weinberg equilibrium. Howerer, population SW and LD with the same waters nearby, the existence of seed flow between the two populations, the higher observed heterozygosity Ho and small inbreeding coefficient FIS, suggesting that water helps gene flow among populations, decreased inbreeding depression within populations and genetic differentiation among populations. The recent establishment of hydropower in region Mount. Emei, with habitat destruction seriously, is bound to inestimable impact of the proliferation of C.omeiensis. Based on the above analysis, the authors made C.omeiensis wild germplasm protection strategy, including protection of Mount. Emei Nature Reserve, establishment of germplasm banks and strengthening the legal protection.