Isolation and characterization of 19 polymorphic microsatellite loci for Atalantia buxifolia (Rutaceae), a traditional m
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MICROSATELLITE LETTERS
Isolation and characterization of 19 polymorphic microsatellite loci for Atalantia buxifolia (Rutaceae), a traditional medicinal plant Miaomiao Shi • Xiaoming Guo • Yizhang Chen Lianxuan Zhou • Dianxiang Zhang
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Received: 9 January 2014 / Accepted: 13 January 2014 Springer Science+Business Media Dordrecht 2014
Abstract To evaluate genetic diversity and structure of a traditional medicinal plant Atalantia buxifolia, 19 polymorphic microsatellite loci were developed and amplified in four multiplex and two individual reactions. The characteristics of these markers were determined in 64 individuals from four sites in South China. Per locus 4–21 alleles were detected. Observed and expected heterozygosities ranged from 0.103 to 0.921 and from 0.483 to 0.934, respectively. These markers are useful tools for future population genetic studies and conservation of genetic resources of A. buxifolia. Keywords Atalantia buxifolia Microsatellites Genetic diversity Medicinal plant Conservation Atalantia buxifolia (Poir) Oliv. (Rutaceae) is a spinous shrub, bearing white fragrant flowers and small globose fruits. It is commonly distributed in forests or thickets near ocean in provinces of South China, as well as in Malaysia, Philippines and Vietnam (Zhang et al. 2008). The species is used as a folk medicine for the treatment of malaria, chronic rheumatism, paralysis, and snakebites, as the extracts from its branch, root or leaves possess various bioactivities such as antifeedant, cytotoxic and antibacterial activities (Yang et al. 2012). In addition, A. buxifolia is
M. Shi X. Guo Y. Chen L. Zhou D. Zhang (&) Key Laboratory of Plant Resources Conservation and Sustainable Utilization, South China Botanical Garden, Chinese Academy of Sciences, Xingke Rd. 723, Guangzhou 510650, China e-mail: [email protected] X. Guo Y. Chen University of Chinese Academy of Sciences, Beijing 10049, China
considered as a close relative of Citrus L. (Bayer et al. 2009), thus it is valued as rootstocks for grafting commercially grown Citrus fruit trees. However, due to frequent human activities, such as over-exploitation and habitat destruction, the populations of A. buxifolia are being destroyed. Assessment of genetic diversity by molecular markers is important for efficient management and conservation of plant genetic resources. Here, we report the isolation and characterization of the first set of polymorphic microsatellite markers for A. buxifolia and expect them to provide a useful tool for population genetic studies. Genomic DNA was extracted from silica gel-dried leaf tissue of one individual of A. buxifolia following the cetyltrimethyl ammonium bromide (CTAB) method (Doyle 1991). Microsatellite loci were isolated using the Fast Isolation by AFLP of Sequences COntaining repeats (FIASCO) protocol from an enriched (TG)n (Zane et al. 2002). Positive clones were sequenced and primer pairs were designed using Primer3web version 4.0.0 (http://pri mer3.ut.ee/). After testing on agarose gel and polyacrylami
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