Development Of EST-SSR Markers For Polyporus Umbellatus And Genetic Diversity

Chuling [Polyporus umbellatus (Pers.) Fries] is a famous traditional Chinese medicinalfungus that belongs to Polyporaceae. Its dried sclerotium has been used as a diuretic intraditional Chinese medicine for centuries. Recently, the sclerotium has been reported toenhance immunity, anticancer, and other pharmaceutical functions, causing a sharp demand inthe market.Polyporus artificial cultivation technology is not mature, and its growth restricted bymany natural conditions, so Polyporus herbs on the market mainly from the wild excavation,resulting in excessive excavation of wild resources, depleting the verge of extinction.Therefore, study on the genetic diversity and phylogenetic relationship of Polyporusgermplasm resources, will has important guiding significantce for innovative use andeffective protection of Polyporus germplasm.EST-SSR makers are associated with the functional genes. It has broad applicationprospects in the genetic diversity and gene maping, genetic and physical map, comparativegenomics and so on. Currently, the EST sequences of Polyporus are limited, development andapplication of SSR markers has not been reported, need for other species SSR markers inPolyporus generic research. Establishment of transferable EST-SSR markers among differentspecies can greatly reduce the cost of primer development，and accelerate the development ofan effective transferable primers.This study begins from Polyporales ESTs, design anddevelop EST-SSR primers, in different producing area of Polyporus DNA as a template forPCR amplification, explores Polyporales in EST-SSR primers transferability, and theapplication research of Polyporus genetic diversity. This study main results are as follow:1Use of Polyporales ESTs from NCBI, development EST-SSR primers.To downloadthe initial segments of ESTs after removal of low quality and redundant sequence processingafter97134no redundant sequences, In total,9520SSRs was obtained in8531ESTs, SSRfrenquence is9.8%. Is given priority to with single nucleotide repeats (85.92%),dinucleotides and Trinucleotides were2.73%and8.57%, respectively. At the same time theEST sequence of Polyporales subordinate branch of SSR has carried on the statisticaldistribution respectively. 2Using the detection of ESTs design SSR primers,30pairs with higher score werechosen. Then these makers were used to PCR amplification the13Chuling germplasm.Twenty-three pairs showed the amplification, accounting for77%of designed primers，and21pairs showed polymorphism, accounting for91%of primers available. EST-SSR primersfrom mononucleotides showed lowest polymorphism.321pairs polymorphism primers were used to analyse the genetic diversity within13chuling germplasm, Genetic similarity coefficien was obtained, the range between0.4222~0.8000.11strains (henan funiu mountain) and2strains (shaanxi taibai mountain north slope)the similarity coefficient of minimum,1strain (shaanxi taibai south slope) and5strains(heilongjiang) has the highest similarity coefficient of0.8000.4Based on the genetic similarity coefficient, using the method of weighted averagemethod to cluster analysis of13species, with similarity coefficient of0.39for threshold,13of chuling germplasm can be divided into three groups, Among them the second class and canbe divided into two subclasses. UPGM clustering tree and principal coordinates analysisresults have not clearly shows the relationship between the geographical distribution and thestrain differentiation.