Individual spatial aggregation correlates with between--popualtion variation in fine-scale genetic structure of Silene ciliata (Caryophyllaceae). Heredity

In August and September 2010, we established one 10x10m plot in each of five S. ciliata populations distributed along the Sierra de Guadarrama (Figure 1, Table 1). Additional information about the selected populations can be found in Lara-Romero et al. 2014. In each study plot, we mapped every adult S. ciliata individual using two high-resolution Differential Global Navigation Satellite System (DGNSS) receivers (Viva GS15, Leica, Switzerland) with an absolute accuracy of 5 cm for x and y coordinates. We also collected leaf material of 96 adult plants per plot for genetic analysis. In order to increase sample size of shorter inter-individual distances for autocorrelation analysis, plants were selected using a clustered random sampling design (sensu Storfer et al. 2007). Namely we randomly selected 24 sample locations in each plot, where we collected leaf material of four individuals located in close proximity to each other. Portions of leaves with no sign of parasites, fungal infection or drought injuries were collected, and subsequently cleaned and dried in silica gel. DNA was extracted using the DNeasy Plant minikit (QIAGEN, Barcelona, Spain) with 10¿20 mg of dried S. ciliata tissue. Ten microsatellite loci previously used in genetic studies of S. ciliata were selected for genotyping: Sci1224, Sci1208, Sci0106, Sci1443 EST-2HTS, EST-8HTS, EST-37HTS, EST-X4-3, EST-G34D06 and EST-G47A02 (García-Fernández et al. 2012b) Specifications of PCR reactions and amplification are detailed in García Fernández et al. (2012b). Samples were run on an automated DNA sequencer (ABI PRISM 3730, Applied Biosystems, California, USA) in Parque Científico de Madrid (Madrid, Spain). Fragment sizes were assigned to alleles using GeneMarker version 1.85 (SoftGenetics, State College, Pennsylvania, USA. After checking, we discarded locus Sci1443 and EST-SSR X4-3 because of inconsistent allelic scoring and the possible presence of null alleles, respectively.