title: Development of Cymbidium ensifolium genic-SSR markers and their utility in genetic diversity and population structure analysis in cymbidiums.
abstract: BACKGROUND
  Cymbidium is a genus of 68 species in the orchid family, with extremely high ornamental value. Marker-assisted selection has proven to be an effective strategy in accelerating plant breeding for many plant species. Analysis of cymbidiums genetic background by molecular markers can be of great value in assisting parental selection and breeding strategy design, however, in plants such as cymbidiums limited genomic resources exist. In order to obtain efficient markers, we deep sequenced the C. ensifolium transcriptome to identify simple sequence repeats derived from gene regions (genic-SSR).
  
  
  RESULTS
  The 7,936 genic-SSR markers were identified. A total of 80 genic-SSRs were selected, and primers were designed according to their flanking sequences. Of the 80 genic-SSR primer sets, 62 were amplified in C. ensifolium successfully, and 55 showed polymorphism when cross-tested among 9 Cymbidium species comprising 59 accessions. Unigenes containing the 62 genic-SSRs were searched against Non-redundant (Nr), Gene Ontology database (GO), eukaryotic orthologous groups (KOGs) and Kyoto Encyclopedia of Genes and Genomes (KEGG) database. The search resulted in 53 matching Nr sequences, of which 39 had GO terms, 18 were assigned to KOGs, and 15 were annotated with KEGG. Genetic diversity and population structure were analyzed based on 55 polymorphic genic-SSR data among 59 accessions. The genetic distance averaged 0.3911, ranging from 0.016 to 0.618. The polymorphic index content (PIC) of 55 polymorphic markers averaged 0.407, ranging from 0.033 to 0.863. A model-based clustering analysis revealed that five genetic groups existed in the collection. Accessions from the same species were typically grouped together; however, C. goeringii accessions did not always form a separate cluster, suggesting that C. goeringii accessions were polyphyletic.
  
  
  CONCLUSIONS
  The genic-SSR identified in this study constitute a set of markers that can be applied across multiple Cymbidium species and used for the evaluation of genetic relationships as well as qualitative and quantitative trait mapping studies. Genic-SSR's coupled with the functional annotations provided by the unigenes will aid in mapping candidate genes of specific function.
authors.affiliation: Zhejiang Academy of Agricultural Sciences, Shiqiao Road 139, Hangzhou, 310021, People's Republic of China. hufanfan1982815@gmail.com.
authors.forename: Xiaobai
authors.initials: X
authors.lastname: Li
authors.affiliation: Hubei University, College of Life Sciences, Wuhan, 430062, People's Republic of China. fengjin1989@yeah.net.
authors.forename: Feng
authors.initials: F
authors.lastname: Jin
authors.affiliation: Zhejiang Academy of Agricultural Sciences, Shiqiao Road 139, Hangzhou, 310021, People's Republic of China. zjulab@163.com.
authors.forename: Liang
authors.initials: L
authors.lastname: Jin
authors.affiliation: USDA-ARS, Dale Bumpers National Rice Research Center, Stuttgart, Arkansas, 72160, USA. Aaron.Jackson@ARS.USDA.GOV.
authors.forename: Aaron
authors.initials: A
authors.lastname: Jackson
authors.affiliation: Agricultural Technology Extension Stations, Shaoxing County Agricultural Bureau, Shaoxing, 312000, Peoples Republic of China. huang680324@163.com.
authors.forename: Cheng
authors.initials: C
authors.lastname: Huang
authors.affiliation: School of Biological Sciences, The University of Hong Kong, Pokfulam Road, Hong Kong, SAR, People's Republic of China. likehu@hku.hk.
authors.forename: Kehu
authors.initials: K
authors.lastname: Li
authors.affiliation: State Key Lab of Rice Biology, International Atomic Energy Agency Collaborating Center, Zhejiang University, Hangzhou, 310029, Peoples Republic of China. shuxl@zju.edu.cn.
authors.forename: Xiaoli
authors.initials: X
authors.lastname: Shu
mesh_terms: D063990:Gene Ontology; D017343:Genes, Plant; D005819:Genetic Markers; D018895:Microsatellite Repeats; D058977:Molecular Sequence Annotation; D029595:Orchidaceae / Q000235:genetics*; D010802:Phylogeny; D011110:Polymorphism, Genetic; D017422:Sequence Analysis, DNA
chemical_list: D005819:Genetic Markers