Cornus Germplasm Resources Survey, Evaluation And Genetic Diversity Study

Cornel(Comus officinalis Sieb.et Zucc.) is a perennial woody plant which belongs to the Cornaceae.The officinal parts are the pericarp of fruit.It is a well-known traditional Chinese medicines and widely used for treatment of vertigo tinnitus,impotence,emission and other symptoms.Cornel plants distribute widely in China.As a dissimilation pollination plant,the germplasms of cornel are diverse in different ecological environments.It brought much difficult to utilize of cornel in production for its unclear germplasm,serious diseases and pests,careless cultivating management,incorrect collection time and method,and nonstandard primary processing etc.Therefore,it is necessary to combine the modern techniques to study,explore,protect and evaluate the germplasm resources of cornel.Taking the cultivar and wild germplasms of cornel from Henan,Shaanxi and Zhejiang as materials and referring to the national germplasm description standard,we established the description criterion,data standard and germplasm description table.Meanwhile,we also established the ISSR molecular marker system of different cornel germplasms and evaluated the germplasms from the morphological traits,the contents of active components and DNA level.This may offer some references to cornel evaluation,good agritulture practice and breeding.The main results are as follows:(1) Based on the national germplasm description standard,we have established the standard of cornel.And then,an in-depth and detailed investigation of germplasm resources of cornel including factors of geography,environment,peculiarities and output has been carried out in Henan province, Shannxi province and Zhejiang province of China.A total of 100 cornel germplasms have been collected and half of them have been submitted to Chinese Medicinal Plants Germplasm Resource Information System.Most of these germplasms were deposited in the germplasm nurseries in Foping, Shaanxi Province.The problems of resource utilization and sustainable development were also proposed.(2) Chacracters of cornel including the colour,shape,length and diameter of fruit,fruit shape coefficient,100-fresh fruit-weight,seed shape coefficient,100-seed-weight,mature phase and contents of ioganin were analyzed.A correlation analysis and cluster analysis were carried out to discuss the diversity and genetic variety of the germplasm resourses of cornel.Results of correlation analysis indicated that the difference among different characters were remarkable.The contents of loganin varied widely,and the coefficient of variation was 29,7%.The coefficient of variation of 100-fresh fruit-weight,fruit length and fruit shape were 21.83%,11.74%and 12.31%,respectively. There were positive correlations among fruit length,the fruit shape coefficient,the 100-fresh fruit-weight,the seed length,and the seed shape.The correlation coefficient between fruit diameter and seed diameter is low,the content of loganin.Other character correlations were not very remarkable.(3) The genetic similarity coefficient of morphological diversity of cornel was 0.538～0.962. With cluster analysis,two groups were clustered by UPGMA phylogenetic analysis.One was the No. 8 germplasm which matured lately,and other 34 germplasms were clustered together.Sub-groups were clustered accordding to the geography originates,the fruit character and the fruit size.(4) The orthogonal design was used to optimize the ISSR-PCR amplification system of cornel. The results showed that 1×PCR buffer,150μmol/L dNTP,1.0μmol/L primer,2.5 mmol/L Mg²⁺, 2.0 U Taq DNA polymerase and 20～80 ng template DNA in a total volume of 20μL were the optimum ISSR-PCR amplification system for cornel.The PCR amplification procedure used in this study was as follow:40 cycles of 30 sec at 94℃,1 minute at 57℃,1 minute at 72℃,with a 10℃minutes pre-inactive of 94℃and a 10 minutes post-elongation of 72℃,and the product was stored at 4℃.(5) Nine primers were seleted from 50 ISSR primers.A total of 179 ISSR bands were obtained by 9 primers,among which 164 were polymorphic bands.The average percentage of polymorphic bands was 91.6%.The genetic similarity coefficient of cornel was between 0.575 and 0.877.By cluster analysis,two groups were clustered by the digital data of polymorphic bands using UPGMA phylogenetic analysis.Twenty germplams of the frist group are coming from Shaanxi Province,and the second group contains germplasms from Shaanxi,Henan and Zhejiang Province.Sub-groups were clustered accordding to the geography originate,the fruit character and the fruit size.The molecular mark cluster results presented some geographical characteristic.