Dogwood Germplasm Genetic Diversity Analysis And The Method Of Building The Core Germplasm Research

Cornus officinalis which has been used generally to subdue blood sugar, treat germs, treat cancer, treat disease of cardiovascular system, prevention and cure AIDS and strengthen immunization and so on, is a tradition Chinese crude drug. It is an important component of many kinds of Chinese patent medicine and clinical formulary. Furthermore, its fruit has been used to develop health food and drink, and its vegetation used to develop afforestation and beautify the landscape. However, it leads to uneven in quality and sell seconds at best quality prices in medicine market for C.officinalis, because of starting late to study and research germplasm resources, lagging behind to collect, preserve, evaluation studies for C. officinalis, and lacking of management experience and cultivar consciousness to pell-mell plant for farmers. Therefore, in order to avoid or abate destruction for tradition medicinal germplasm resources of C.officinalis, protect its genetic resources and xenogeneic diversity, develop and utilize well germplasm resources of C. officinalis, we must be badly in need of solve the above-mentioned problem from germplasm.In this paper, the genetic diversity of C.officinals, which was collected from Zhejiang, Henan and Shaanxi provinces in China were analyzed on DNA molecular level by ISSR molecular marker method.The construction core collection methods can be analyzed based on it. At the same time, this paper analyzed the content of loganin of C.officinals from three provinces in China by HPLC. It can provide accordance for evaluation and sieving good species. The main results of the study are as follows:1. The high quality genomic DNA of C.officinals, is suitable for ISSR analysis, was extracted by an improved method of CTAB. The ISSR reaction system for C.officinals was established, screened18primers amplification products are clear, stable and high polymorphism from48primers based on the pretty ISSR reaction system. Those18primers were determined the optimal annealing temperature to utilize annealing temperature gradient experiments. Screened18primers amplified256site, the total233polymorphic site were amplified and the polymorphic loci was91.02%, the results about Na,Ne,H and I were1.9102,1.4847,0.2928and0.4459. The results showed that the C.officinals germplasm resource had a higher genetic diversity and representativeness on DNA molecular level, relatively higher genetic diversity from Zhejiang and Henan, and lowest genetic diversity from Shaanxi. There were some gene flow(Nm=2.1740) among three main producing areas of C.officinals. Furthermore, genetic variation within populations is major. Genetic relationships were close C.officinals germplasm resource from Zhejiang and Henan areas, and farther C.officinals germplasm resource from Shaanxi areas.2. The results about three sampling technique (CG、SC and LDSS) and seven sampling ratio (40%,35%,30%,25%,20%,15%,10%) showed that the LDSS and25%sampling ratio were the most well suitable method for constructing the C.officinals core germplasm by analysis of genetic diversity parameter from the above-mentioned amongst samples. The final core collection numbers (18) were only24.66%numbers of primary collection (73), but the Nei’s gene diversity (H) and Shannon’s Information index (l), which were higher than that of the primary germplasm resources, were0.2937and0.4427, respectively. The other genetic diversity parameters, the percentage of polymorphisms loci (Ap), observed number of alleles (Na) and effective number of alleles (Ne) were remained94.85%,97.55%and99.28%of the primary germplasm resources. The genetic distance (D) of core collection (0.2320～0.6854) increased compared to the genetic distance (D) of initial germplasm (0.0984～0.6776), it showed that redundant material of the initial germplasm resources were effectively removed, and cutting down the genetic redundancy of the initial germplasm resources. They have T-test between the H,(0.2948),HS (0.2102), GST (0.2869),Nm (1.2428) of the core collection and the H,(0.2957),HS (0.2404), GST (0.1870),Nm (2.1740) of the initial collection, their difference were not significant (P=0.435>0.05).It showed that their hereditary constitution are identical both the core collection and the initial collection, and got the aim that the genetic resource were kept to be maximize and high efficiency, but germplasm numbers were smallest.3. In accordance with 《Pharmacopoeia of the People’s Republic of China》,this paper determined the active constituent of58pieces of C.officinals from the three main producing areas, the contents of loganin. The results showed that the mean value was0.72%that exceeded Pharmacopoeia approved (0.6%). The mean contents of loganin in Henan area is the first (0.983%), Zhejiang (0.816%) secondary, and Shaanxi was the lowest (0.546%). The ANOVA showed that it was very significance to the contents of loganin amongst populations (P=0.000<0.01). Cluster analysis results showed that genetic distance on content of loganin of Cornus officinalis is nearer between Henan and Zhejiang, and is farther between Henan and Shaanxi.The population clustering’s result on content of loganin and the result by ISSR markers are the same. It showed that the DNA molecular level and the active constituent level have a higher consistency among three main producing areas of C.officinals.