| Loropetalum subcordatum(Benth.) Oliv.(Hamamelidaceae) is evergreen trees or shrubs; leaves alternate, leaf blade ovate or elliptic, tender leaf is red, elder leaf is dark green. Flower is white, 5 petals. This species has a narrow distribution and is endemic to China. It has been categorized as a National Protected Species(Class II) and listed in International Union for Conservation of Nature. Loropetalum subcordatum is very important and meaningful for the research of Ecology diversity, Species diversity, Genetic diversity and Plant evolution. At present, Loropetalum subcordatums' habitats has been suffered from damaged and degraded, even extincted, due to frequent human activities. Therefore, it is extremely urgent to protect Loropetalum subcordatum. Realizing the genetic diversity and genetic structure of endangered species is the foundation and precondition to protect those plants successfully. This study analyzed genetic diversity of 6 nature and 1 ex situ populations of Loropetalum subcordatum by using SRAP molecular markers. And coming up with conservation suggestions, supplying a reference and establishing theoretical basis for ex situ conservation and reintroduction of Loropetalum subcordatum through analyzing and discussing experimental data. The major research results of this study are: 1. The establishment SRAP-PCR reaction system of Loropetalum subcordatumThe single factor experiment of 20?l SRAP-PCR reaction system of Loropetalum subcordatum results show that optimal amount of templates DNA is 40~80ng, amount of Ex Taq 1.5~3.0U, d NTPs concentration 0.25~0.4mmol/L, Mg2+ concentration 2.00~3.50mmol/L and primers concentration 0.2~0.35?mol/L. The orthogonal experimentof 20?l SRAP-PCR reaction system of Loropetalum subcordatum results show that the best 20?l reaction system is amount of templates DNA is 60 ng, amount of Ex Taq 2.5U, d NTPs concentration 0.25mmol/L, Mg2+ concentration 3.5mmol/L and primers concentration 0.3?mol/L. 2. Genetic diversity of nature populations of Loropetalum subcordatumAmplification from the fifteen primer combinations for all the accessions produced 553 fragments in total, and a total of 359 bands displayed polymorphism with the PPF of 63.28%. Among six nature populations, GZ exhibited the highest genetic diversity with PPF of 21.7% and DY showed the lowest genetic diversity with PPF of 3.98%, GX, WG, BY and ZL was 17%, 13.2%, 7.78% and 7.23%, respectively. The amount of observed alleles(Na) was 1.0398~1.2170, the effective number of alleles(Ne) was 1.0247~1.1620, Nei's genetic diversity(H) was 0.0145~0.0904, Shannon diversity(SI) was 0.0217~0.1308 among six nature populations, respectively. The total gene diversity(Ht) of Loropetalum subcordatum was 0.2217 and the average gene diversity within population(Hs) is 0.0469. Genetic divergence among populations(Gst) was 0.7884, P<0.001, which indicated that there are fewer gene exchange had taken place among nature populations. Those results revealed that Loropetalum subcordatum had a low genetic diversity. 3. Genetic structure of nature populations of Loropetalum subcordatumLoropetalum subcordatum had high genetic divergence, because gene flow among populations was very low(Nm*=0.1342), and genetic differentiation among population was very high(Gst=0.7884), which suggested that 78.84% of genetic variation occured among populations.The result of the cluster analysis by UPGMA, principal coordinate analysis and Bayesian algorithm to analyzed the genetic structure of nature population of Loropetalum subcordatum was the same, which showed that six nature population was categorized as three groups. The first group included GZ and GX population, and the second group only had WG, the last group contained BY, ZL and DY. The result of Mantel test showed that there are significant correlation between genetic distance and geographical distance(r=0.91, p=0.01), and linear equation(y=1740.3x-68.095) revealed that this is positive a correlation. 4. Genetic diversity and structure of ex situ population of Loropetalum subcordatumGenetic diversity of Zhongshan ex situ population(ZS) of Loropetalum subcordatum(PPF=7.59%) was lower than Genetic diversity of Wugui Mountain nature population(WG) of Loropetalum subcordatum(PPF=13.2%).There are fewer genetic variation(Gst=0.0889) occurred between ex situ population of Loropetalum subcordatum(ZS) and its provenance(WG). The amount of observed alleles(Na) and the effective number of alleles(Ne) of ZS was similar with WG, the difference between two populations just only within 5%. But the polymorphism bands, Nei's genetic diversity(H) and Shannon diversity(SI) of ZS, just accounted for 57.53%, 62.42% and 61.57% of WG respectively were far lower than WG. Those results indicated that genetic diversity and structure of ZS did not covered its provenance(WG) and this difference was very huge between those two populations. Finally, confirming 14 individuals of GD can cover its all the genetic diversity by using POPGEN 32. |
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