Genetic Diversity In O.japonicus (L.F.) Ker-Gawl. Based On RAPD Analysis And Filtration Of Differential Marker

The use of the RAPD technique was investigated on 16 natural populations of O.japonicus (L.f.) Ker-Gawl. in Sichuan, Hubei, Zhejiang, Shanghai, and two wild relational species, O. bodinieri Levl. and O.bockianus Diels, which belong to the same genus with O.japonicus (L.f) Ker-Gawl., after optimizing PCR conditions. Due to high sensitivity of RAPD analysis to reaction conditions, main factors affecting the results including concentrations of Mg2+, Taq DNA ploymerase, primers and templates, and number of PCR cycle etc., were examined. Conditions applicable to RAPD analysis of O.japonicus were determined. The screening of 40 decamer oligonucleotides allowed the selection of 8 primers used for the analysis. A total of 504 RAPD markers were produced from the 8 selected primers. Fingerprints of 18 samples had been constituted. Relationships among the 18 populations were clustered using the least distance method analysis in a dendrogram. The largest distance was found between O.bockianus Diels, O. bodinieri Levl. and O.japonicus (L.f.) Ker-Gawl.. There had no difference in four samples of Mianyang City during dusting. It investigated their special characters were related to geographic regions. The largest distance was found between big leaf type population in SichuanUniversity and the other populations. Large genetic variation occurred among the different clime and leaf-type populations.To establish a kind of rapid and efficacious molecular method to identify the Ophiopogon japonicus(L.f.)Ker-Gawl. by officinal directly, we prepare DNA from its dried root for RAPD. Extracted total DNA from dried root of the sprawled sample of Mianyang Yingcheng. Same 40 random decarner primers were screened for RAPD fragments.il identifiable primers were selected. Extracted DNA from leaves dried through silica gel and fresh leaves for contrastive experiment in the same time. In result, there were no any obvious different RAPD results in the three kinds of template DNA from electrophoresis. It was concluded that the quality and quantity of the template DNA prepared from officinal was enough for RAPD.Research of transferring RAPD marker to SCAR marker also had been done. Select two specific fragments to clone and sequence. Then design specific primers to amplify the fragments as verification. Though the verification had not been completed, experience for advancing works had been accumulated.