Genetic Diversity Research On Endangered Tibetan Medicine Sinopodophyllum Hexandrum(Royle)Ying And Anisodustanguticus

Studying Genetic diversity on the Genuine Medicinal Materials Sinopodophyllum hexandrum(Royle)Ying and Anisodustanguticus(Maxim.Pascher)in edge-region of qinghai-tibet plateau of north Gansu Province,China.Sinopodophyllum hexandrum(Royle)Ying is a more commonly used Tibetan medicine,and also a rare medicinal plant in China.It belongs to the endangered population and is a national class Ⅱ protected plant.The Anisodus tanguticus(Maxim.Pascher)is a perennial herb with the alias Zhang Liushen,Tibetan osmanthus,Zhang Liu(Qinghai),and is also one of the more commonly used Tibetan medicines in China.It is listed as a national class II protected plant and is biologically diverse Internationally protected species.This study collected the wild genetic resources of Sinopodophyllum hexandrum(Royle)Ying and Anisodus tanguticus,and extracted genomic DNA,optimized ISSR primer selection and amplification system,and optimized the ISSR-PCR reaction system for the molecular structure of Sinopodophyllum hexandrum(Royle)Ying and Anisodus tanguticus.To test the genetic diversity of two endangered wild medicinal plants of Sinopodophyllum hexandrum(Royle)Ying and Anisodus tanguticus,and to analyze the genetic diversity of their wild populations,so as to propose the conservation and sustainable use of each Related suggestions of them.The main findings are as follows:This research covers the Sinopodophyllum hexandrum(Royle)Ying and Anisodus tanguticus,collecting with the wild heredity resources and withdraw genome DNA,carry on ISSR to lead thing sieving and expand system to build up excellent turn,respond system through excellent turned ISSR-PCR to the Sinopodophyllum hexandrum(Royle)Ying and Anisodus tanguticus,marking experiment and record an experiment result to combine analytical wild reside the heredity of cluster of genetic diversity.Main research result as follows:(1)Optimizing the ISSR-PCR system.Analyzing the results of PCR by optimizing five major factors(template,Mg2+,Taq DNA enzyme,dNTPs and primer)of ISSR-PCR reaction in four levels through the combination of orthogonal experimental design and signal factor ananlysis..The result of Experimental design is that primer>Taq DNA enzyme>template>dNTPs>Mg2+,The best system is established as follows:2.5 mmol/L Mg2+,2.5 mmol/L,0.2 mol/L dNTP,0.4 μmol/L primer,0.5 U Taq DNA polymerase,40 ng DNA,2.5 μL 10× Buffer.The system has high stability,abundant polymorphism characteristics.(2)Genetic diversity analyzing of wild Sinopodophyllum hexandrum(Royle)Ying.The amplifying products from 153DNA based on 11 ISSR primers technique,Total 155 alleles were amplified.the average percentage of polymorphic loci was 99.2%.The total genetic diversity(Ht)of the 13 populations of 13 wild peaches was 0.3227,and the genetic diversity(Hs)within the population was 0.2101,indicating that the genetic diversity was from among the populations.The gene differentiation coefficient(Gst)was 0.3489,and the genetic differentiation between species was 34.89%.The remaining 65.11%of the genetic differentiation occurred within the population and was significantly higher than that between the populations.The differentiation is big.Gene flow(Nm)= 0.9331,indicating that the gene flow within the population is very abundant.The results showed that among the 13 wild peaches,7 populations had high genetic diversity.There was no correlation between genetic distance and geographical distance among the populations.There were high polymorphisms and abundant diversity among them.(3)Genetic diversity analyzing of wild Anisodus tanguticus,.The amplifying products from 127 DNA based on ISSR makers technique were used to analyze the genetic diversity of 11 populations of wild Anisodus tanguticus.Total 131 alleles were amplified by using 10 ISSR primers.The number of alleles in per primer range was 11～15 with an average of 13.1,and 858 polymorphic loci were got finally.The average percentage of polymorphic loci(PPB)was 59.54%.The Nei’s gene diversity(H)and Shannon’s information index(I)were 0.220 4 and 0.325 4 respectively.The genetic distances range was 0.073 3～0.306 2,and Mantel test P = 0.002.The genetic polymorphism among 11 populations of Anisodus tanguticus is relatively abundant and there is a directly relationship between genetic distance and geographical distribution among them.(4)Based on the results and comprehensive genetic diversity analysis,That in addition to adopting an on-site protection strategy to protect the genetic diversity of its original population,the wild genetic resources of Sinopodophyllum hexandrum(Royle)Ying should be focused on the ex situ conservation of different germplasm resource,Keeping stability of their gene flow with the No.2 Gansu Luqu Shilin population that as the in-situ core protection site.No.3 Wagougou population and No.7 Xinglong Shanxishan Populations are focued on observation and detection.Protecting all the genetic resources of Sinopodophyllum hexandrum(Royle)Ying.At the same time,germplasm resources of Anisodus tanguticus from different regions are used for exsit conservation and breakthrough in geographic isolation(including rivers,mountains,and artificial roads).Ditches and other factors that hinder gene exchange among populations and ensure the flow of genes.