Studies On Genetic Relationship Of Culinary Rhubarb And Official Rheum Based On ISSR And SRAP Markers

Culinary rhubarb is the perennial herbaceous plant,which has thick and juicy petioles.It is the ideal health care vegetable which contains many nutrients,such as cellolose A,vitamin C,calcium,phosphorus and succinic acid.In Europe,North Americal and Oceania,culinary rhubarb has been used for commercial cultivation.In the 17 th century,the Chinese medicinal rhubarb was introduced to Europe,after a long period of natural,it evolved into culinary rhubarb.However,the relationship of culinary rhubarb and official rheum is not reported.This paper studied seed morphology of twenty-two varieties and ISSR and SRAP molecular markers analysis.The optimization of ISSR-PCR reaction system was conducted and primers screening and genetic relationship were analysed between culinary rhubarb and official rheum.This study achieved the following main results:(1)The optimal ISSR-PCR reaction system was established for culinary rhubarb,e.g in a 25 μL reaction system contained 10×PCR buffer 2.50 μL,template DNA 125 ng,Mg2+ 2.50 mmol/L,d NTPs 0.35 mmol/L,primer 0.50μmol/L,Taq DNA polymerase dosage 0.04 U/μL,with residual volume supplied by double distilled water.(2)Through the investigation of twenty-two varieties of rhubarb agronomic traits were cluster analysied by SAS v9.2 software from different stains(including seed length,seed width,seed wing broadth and thousand seed weight).Twenty-two copies of materials can be divided into four classes when the maximum distance was 3.5.In addition,the clustering analysis showed that there was a bigger difference between RT and other 21 materials in seed character.(3)Using the variety of Culinary rhubarb R34 and variety of official Rheum SC respectively to screen 100 ISSR primers and 182 SRAP primer combinations.The results showed 13 ISSR primers and 32 SRAP primer combinations of distinct and rich in polymorphism.In addition,ISSR-PCR amplification received 74 allele loci,in which 59 is polymorphism loci.SRAP-PCR amplification for a total of 227 polymorphic bands,containing 206 polymorphic.Those results show that these two kinds of molecular markers can provide abundant genetic information and can be used to analyze the rhubarb genetic diversity.(4)The genetic similarity and genetic distance of 22 rhubarb materials were analyed by using POPGENE1.32 software.Genetic similarity based on ISSR markers ranged from 0.3220-0.8983,the genetic distance ranged from0.1072-0.9865,the result showed the genetic similarity coefficient between rhubarb SC and YN is the largest,indicating that these two varieties is the most close.The coefficient of ‘tudahuang’ and RG is the least,indicating that these two varieties is the most distant.Genetic similarity based on SRAP markers ranged from 0.3301-0.8495.The result showed the genetic similarity coefficient between rhubarb RG and RO is the largest,indicating these two varieties is the most close.The coefficient of ‘nandahuang’ and Ro is the least,indicating these two varieties is the most distant.(5)The phylogenetic tree of 22 rhubarb samples were constructed by NTSYS pc2.10 c.Based on ISSR and SRAP markers,the 22 samples could be divided into four groups when genetic similarity coefficient was 0.60.R54,R34 and RO were belonged to a group,‘tudahuang’ was individually in a group.‘chuandahuang’,‘kudahuang’,‘maodahuang’ R19,SC,ZJ,HN,GZ and HB were belonged a group.The left 9 samples were belonged to a group.