A reduced representation libraries approach for nuclear marker development via 454 sequencing applied on Tetramorium (Hymenoptera: Formicidae). Tetramorium

To clarify species boundaries in the cryptic Tetramorium caespitum/impurum complex, a multi-discipline approach is necessary. Nuclear DNA markers constitute one basal step in species delimitation. We established a reduced representation library (RRL) based on amplified fragment-length polymorphism (AFLP) PCR products of four species. Amplicons were size-selected by electrophoresis, DNA recovered from the agarose gels, and 454 libraries construced by a commercial provider. Pyrosequencing yielded 490,155 reads, from which we designed cross-species amplifying primer pairs for 28 loci. Wetlab procedures, bioinformatic pipeline, alignment statistics, and the developed primers are presented.