Genomic DNA enriched for microsatellites: 14 Conifer Species
Genomic DNA from diploid needle tissues were isolated from all trees, and genomic libraries were constructed using standard Illumina library construction, but with in-line 4 base pair barcode adapters described in Cronn et al., 2008 (Nucleic Acids Res. 36(19):e122. DOI:10.1093/nar/gkn502 ). To enrich microsatellite loci for these conifer genomes, we enriched genomic DNA libraries for di- and tri-nucleotide repeats using the procedure outlined in Jennings et al., 2011 (Mol Ecol Resour. 11(6):1060-7. DOI: 10.1111/j.1755-0998.2011.03033.x).  The resulting 14 enriched libraries were pooled to equimolar representation library, and 5 pmol of the library was sequenced on an Illumina Genome Analyzer II using 80 bp paired end reads (University of Oregon Genomics Core Facility, Eugene, OR, USA).  Individual taxon libraries were identified and sorted by barcode using a custom Perl script (https://github.com/listonlab/utility-scripts/blob/master/bcsort_v5.pl), and individual barcodes include: 5-CACT,  Pinus albicaulis; 5-GCTT, Pinus coulteri; 5-CTGT, Pinus.flexilis; 5- TTGT, Pinus glabra; 5- GATT, Pinus jeffreyi; 5- TGCT, Pinus monticola; 5- ATGT, Pinus palustris; 5- CAGT, Pinus radiata; 5- CCCT, Pinus sabiniana; 5- GTAT, Picea glauca; 5- GGGT, Chamaecyparis lawsoniana; 5- CGAT, Tsuga canadensis; 5- AACT, Abies concolor; 5- ACGT, Abies lasiocarpa.Since the sequencing read length of this early-generation Illumina sequencing is too short to span complete inserts, a strategy to identify microsatellites from partial sequences was devised; this can be referenced in Jennings et al., 2011.