Versatility. Of Dendrobium Candidum Chloroplast Microsatellite Development And Application Of Its Kinds

As a rare and endangered perennial herb endemic to China, Dendrobium officinale Kimura et Migo (Orchidaceae) is famous for its unparalleled medicinal value and ranked "the first of the Chinese nine kinds of supernatural medicinal herbs". Unfortunately, the natural resource of D. officinale has been in severe danger of extinction because of habitat deterioration and human exploitation enticed by high profit using the stems as good traditional Chinese medicine in recent years.Microsatellites or simple sequence repeats (SSR), a group of tandem repeated sequences constituted from mono-, di-, tri-, tetra-, penta-, or hexa-nucleotide units and widely distributed throughout nuclear and mitochondrial genome as well as chloroplast genome in eukaryotes. To date, chloroplast microsatellite markers (cpSSR) have been developed for a variety of species and demonstrated utility in studying evolution, differentiation, genetic diversity and genetic structure of plant population.In this study, nine pairs of polymorphic chloroplast microsatellite primers were developed for D. officinale. Levels of polymorphism were tested across a total of 55 individuals from four natural populations. The genetic differentiation was characterized using the analysis of molecular variance. Allele numbers varied from two to four per locus, while the gene diversity ranged between 0.133 and 0.703. The number of haplotypes ranged from four to six per population. The proportion of genetic differentiation within population was higher than that among populations. These polymorphic cpSSR markers developed for D. officinale will be a useful tool to investigate genetic diversity, population genetic structure, evolution of D. officinale and establish effective conservation strategies.Transferability of the nine polymorphic chloroplast microsatellite primers was checked on a set of 51 Dendrobium individuals belonging to 17 different species, and the level of polymorphism of these markers across Dendrobium moniliforme (L.) Sw. and Dendrobium loddigesii Rolfe was examined, as well. The genetic differentiation was characterized using the analysis of molecular variance. Three markers could be transferable to all the species tested, while the remaining six markers successfully cross-amplified in most species tested. All of them were polymorphic in D. moniliforme, while seven of which were polymorphic in D. loddigesii. In three populations of D. moniliforme, allele numbers per polymorphic locus and the gene diversity ranged from 2 to 4 and 0.167 to 0.582, respectively. The number of haplotypes ranged from four to six per population. In four populations of D. loddigesii, allele numbers per polymorphic locus and the gene diversity ranged from 2 to 3 and 0.167 to 0.718 separately. The number of haplotypes ranged from two to five per population. The same as the D. officinale, the proportion of genetic differentiation within population of two species was higher than that among populations. Besides, cross-amplification experiment showed that these cpSSR loci may be commonly applicable to other Dendrobium species.