Abstract
Premise Camellia reticulata, which is native to southwestern China, is an economically important plant belonging to the family Theaceae. We developed expressed sequence tag–simple sequence repeat (EST‐SSR) markers for C. reticulata, which can be used to investigate its genetic diversity, population structure, and evolutionary history.Methods and ResultsWe detected 4780 SSRs in C. reticulata from Camellia RNA‐Seq data deposited in the National Center for Biotechnology Information's expressed sequence tags database (dbEST). Primer pairs for 70 SSR loci were designed and used for PCR amplification using 90 individuals from four populations of C. reticulata. Of these loci, 50 microsatellite markers were successfully identified, including 11 polymorphic markers. The allele number per locus ranged from two to seven (mean = 4.182), and the levels of observed and expected heterozygosity ranged from 0.044 to 0.567 and from 0.166 to 0.642, respectively. Eleven primer pairs amplified PCR products in three other species of Camellia (C. saluenensis, C. pitardii, and C. yunnanensis).ConclusionsThe set of microsatellite markers developed here can be used to study the genetic variation and population structure of C. reticulata and related species and thereby help to develop conservation strategies for this species.
Highlights
PREMISE: Camellia reticulata, which is native to southwestern China, is an economically important plant belonging to the family Theaceae
We detected 4780 SSRs in C. reticulata from Camellia RNA-Seq data deposited in the National Center for Biotechnology Information's expressed sequence tags database
Camellia reticulata Lindl. (Theaceae) is an economically important ornamental flowering shrub or small tree that grows in Yunnan Province, southwestern Sichuan Province, and western Guizhou Province of China (Ming et al, 2000)
Summary
Fresh healthy leaves collected from 90 individuals of C. reticulata sampled from four wild populations from Yunnan Province, China, were freeze-dried or silica-dried. Clean EST sequences were clustered and assembled into contigs and singletons using CAP3 (Huang and Madan, 1999), generating 17,989 unigenes consisting of 5099 contigs and 12,890 singletons. These unique sequences were further used to screen for the presence of microsatellites using the MISA Perl program (Thiel et al, 2003). We randomly selected 70 primer pairs and tested them for PCR amplification in 12 accessions of C. reticulata (three individuals in each population, Appendix 1) in an initial screening test. Eleven loci showed polymorphisms (Table 1), and 39 loci were monomorphic (Appendix 2) These 11 polymorphic primers were used in 90 individuals of C. reticulata (four populations) for the population genetic analyses using the same protocol as the initial test. Locus CSSR2 CSSR5 CSSR11 CSSR17 CSSR18 CSSR19 CSSR35 CSSR36 CSSR38 CSSR45 CSSR48
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