SRAP and SSR molecular markers of pink flower Dendrobium genetic diversity and population genetic structure
|School||Nanjing Normal University|
|Keywords||Pink flower Dendrobium SRAP SSR Genetic diversity Population Genetic Structure|
Dendrobium loddigesii Rolfe is an endangered epiphytic orchid with extremely high medicinal and ornamental value. The stems of D. loddigesii are widely known as the most precious traditional Chinese medicine, its main function is nourishing yin and clearing away heat-evil, benefiting the stomach, moistening the lung and relieving cough and prolonging life. D. loddigesii is at the risk of extinction because of habitat deterioration, human exploitation enticed by high profits, low germination rate and slow growth. Therefore, it is extremely urgent to determine the level of the genetic diversity and pattern of population genetic structure of an endangered species as it is an important prerequisite for effective conservation and management.In this thesis, sequence-related amplified polymorphism (SRAP) was employed to investigate the genetic diversity and population genetic structure in nine natural populations of D. loddigesii. The orthogonal design was used to optimize SRAP-PCR amplification system on D. loddigesii. Seventeen SRAP primer combinations generated a total of 231 clear amplification bands encompassing 187 (80.95%) polymorphic bands. A high level of genetic diversity was detected (PPB=80.52%, H =0.2743, I=0.4113) at the species level. There was a moderate genetic differentiation (Gst=0.304) among populations. Two main clusters were detected by cluster analysis using the unweighted pair-group method with arithmetic average (UPGMA), ClusterⅠincluded populations (ML, HC, LY, YD, SG) from Guizhou, Guangxi and Guangdong provinces. ClusterⅡincluded populations (BS, HH) from Hainan and Yunnan provinces, which was consistent with the result of principal coordinate analysis (PCA). Mantel test revealed that no significant positive correlation was found between genetic distances and geographic distances (r=0.3138; P>0.05).Secondly, new micro satellite DNA was isolated from D. loddigesii genome DNA through the methods of entichment by magnetic beads. Twelve polymorphic microsatellites were development for D. loddigesii, and their polymorphism was tested across all individuals from nine natural populations. A total of 98 alleles were detected with an average of 8.2 alleles per locus. The average expected heterozygosity and polymorphism information content values were 0.690 and 0.637, respectively. Results showed that these twelve new microsatellites had high polymorphism. They would be useful tools for estimating the genetic diversity and population genetic structure of D. loddigesii. Lastly, twelve novel microsatellite markers were employed to investigate the genetic diversity and population genetic structure of D. loddigesii. A high level of genetic diversity was detected. The expected heterozygosity (He) ranged from 0.484 (BS) to 0.665 (HC), with an average of 0.582. Analysis of molecular variance showed that the main variation component existed within populations (82.02%) rather than among populations (17.98%). A dendrogram cluster analysis resolved the nine populations into two main clusters, which was consistent with the result of principal coordinate analysis (PCA). Mantel test revealed that no significant positive correlation was found between genetic distances and geographic distances (r=0.4126; P>0.05).In conclusion, this is the first study about the genetic diversity and population genetic structure of D. loddigesii by SRAP and SSR. The insights gained from investigating the genetic diversity and population genetic structure of D. loddigesii are important in developing a sound conservation program. In the further, these two kinds of molecular markers would provide important information for forensic identification, genetic mapping, conservation biology and breeding programs of D. loddigesii.