Development and Transfer Analysis of SSR in Dendrobium

To date,only 100SSR markers in Dendrobiumhave been developed,which are far from sufficient for research applications.To develop molecular markers,we mined SSR of Dendrobium from public nucleotide data through bioinformation methods.Some 1343Uni-DNA sequences were assembled from the 3599DNA sequences of Dendrobiumfrom GenBank.By scanning the Uni-DNA sequences,283SSRs were distributed in 205Uni-DNA sequences,with an average frequency of 1SSR per 2815bp.Sequence alignment indicated that 86of the 205SSR-DNA sequences had already been used to design primers.In this study,76primer pairs were designed from the remaining 119sequences for transferability analysis among 32 Dendrobium species.Results showed that 47primer pairs were amplified effectively with transfer rates ranging from 51.1%to 95.7%(average75.9%).Of which,46primer pairs were able to detect polymorphism among the Dendrobium species with 2-8alleles(average 4.0alleles).Ten pairs of polymorphic primers were selected to detect polymorphism in 60accessions of D.officinale,and 2-5alleles(average 3.4alleles)were found per SSR locus.Based on the SSR amplification pattern,the 60accessions of D.officinale were clustered into five clusters,and phenotypes were closer within clusters than between clusters.The sequencing of the amplified fragment of DM121revealed that allele variation within D.officinale was attributed mainly to the variation of SSR repeat numbers,whereas allele variations among Dendrobium species were also caused by a single base indel and substitution in the microsatellite flanking region.