Genetic Diversity Of RhododendronmolleG. Don In China Based On ISSR Analysis

The improved 2×CTAB method was used to extract DNA from RhododendronmolleG.Don plant-leaves which were gathered from Jinxi,Lushan Mountain,Jinggangshan Mountain,Wuning and Richang. The recycling parameters were optimized and optimal reaction system suitable for ISSR-PCR amplification was established in the paper. The primers that can produce polymorphic, clear and bright bands have been selected for 1SSR amplification analysis on 39 samples in RhododendronmolleG.Don species from 167 random primers.The results show that acetone can improve the purity of output. The optimal reaction system of ISSR-PCR for RhododendronmolleG.Don was determined as follows: template DNA 30～45ng, Mg²⁺2.0～2.5mmol/l, dNTPsO.l 0.15mmol/l,primer 0.3～0.4umol/l, Taq DNA polymerase 0.5～1.0U; 10×PCR buffer 2.5μιand ddH2O in total 25μιreaction system. The optical amplification program was obtained: 94°C 5min, lcycle; 94°C lmin, 55°C30s , 72°C2min,30cycle; 72°C lmin,lcycle;4°C stop.13 primers that can produce polymorphic, clear and bright bands have been selected for ISSR amplification analysis on DNA samples in 5 RhododendronmolleG.Don populations from 167 random primers. Using 13 primers, the total 196 loci were found ,190 of which were polymorphic loci. The percentage of polymorphic loci is 96.94%.0,1 matrix constructed according to amplified maps was inputted into computer. The result of POPGENE analysis indicated that the level of genetic diversity of RhododendronmolleG.Don was high (H=0.3342, I=0.5007). Nei's genetic diversity analysis and analysis of molecular variance (AMOVA) show that there was a certain level of genetic differentiation (Gst: 0.5084 and 0.5320) among the five populations. The dendrograms of genetic relationships among populations was constructed based on Nei's genetic distance. JinxI Lushan Mountain,Jinggangshan Mountain and Wuning populations clustered in a clade; Ruichang population was in a clade. According to Mantel, there is a certain correlation between genetic distance and geographic distance, but not observably.Some suggestions were taken through analysis on genetic diversity and genetic structure of RhododendronmolleG.Don: 1. protection on the spot and transplanting species for protection; 2. cross-transplant among populations to enhance the gene flow; 3. protecting and rebuilding suitable environment for RhododendronmolleG.Don populations.