Microsatellite loci for the social wasp Polistes dominulus and their application in other polistine wasps

The social wasps of the genus Polistes are an important model system for understanding the evolution of cooperation. Their relatively simple societies lack the distinct morphological castes which characterize many of the social insects, and newly emerged females possess a variety of reproductive options (Reeve 1991). A female may remain on her natal nest as a helper gaining indirect fitness; usurp a foreign nest and become reproductively dominant; initiate a new nest independently; reproduce on a satellite nest; or initiate a new nest in cooperation with other wasps (Strassmann 1981; Reeve 1991; Mead et al . 1995; Cervo & Lorenzi 1996; Queller et al. 2000). By characterizing the reproductive payoffs associated with different reproductive strategies, we are better able to understand how cooperative societies are maintained. Recently, microsatellite genetic loci have greatly extended our ability to characterize the reproductive strategies used by social wasps (Hughes 1998; Queller et al . 1993a). Using microsatellite loci we can reconstruct pedigrees, and estimate relatedness. Using this information, unobserved events such as queen death, nest usurpation or past reproductive dominance can be inferred (Queller et al . 1993a,b; Field et al . 1998; Hughes 1998). In this paper, I describe microsatellite loci isolated from the social wasp Polistes dominulus , one of the best studied Polistes species. We followed published protocols for the isolation of microsatellite loci (Strassmann et al . 1996) with clarifications and modifications to those protocols as noted below. DNA was extracted from 1 to 1.5 g of pupal thoraces ground in a mortar and pestle which had been chilled in liquid nitrogen. The ground tissue was suspended in grinding buffer (0.1 m NaCl; 0.1 m Tris-HCl, pH 9.1; 0.05 m EDTA; 0.05% SDS), and purified three times with phenol:chloroform:isoamyl alcohol (25:24:1), and then three times with chloroform:isoamyl alcohol (24:1). The purified genomic DNA was then ethanol precipitated, and resuspended in distilled water. Genomic DNA was digested with Sau 3aI, and 300–1000 bp inserts were ligated into the pZErO –2 plasmid (Zero Background cloning kit, Invitrogen) digested with Bam HI. We transformed TOP10 cells (Invitrogen) to obtain approximately 5000–6000 clones. Nylon replicates of the genomic library were probed with five oligonucleotides (AAT 10 , AAG 10 , AAC 10 , TAG 10 , and CAT 10 ) which were end-labelled with [ γ 33 P]-dATP. Probes of the nylon replicates yielded 151 positives and subsequent probing of plasmid DNA on the southern blot confirmed 34 unique positives. Clones which were positive on the southern blot were sequenced on an ABI 377 automated sequencer (Perkin-Elmer), and 19 sets of polymerase chain reaction (PCR) primers were designed from the 28 resulting sequences using Mac Ventor 5.0 (Kodak Scientific Imaging Systems). We optimized the PCR primers on an MJ Research PTC100 thermocycler using 10 μ L reactions (Peters et al . 1998), and assessed within-species polymorphisms for eight species of polistine wasps, using from one to eight unrelated females for each species (Table 1). PCR products were visualized on 6% polyacrylamide/8 m Urea sequencing gels. Twelve of the 19 loci tested were polymorphic within our P. dominulus population and had a mean observed heterozygosity ( H O ) of 0.76. Loci with a minimum of five uninterrupted repeats were polymorphic, and heterozygosity increased logarithmically with the number of uninterrupted repeats (Fig. 1; logarithmic regression, R 2 = 0.454, P = 0.0016). The loci retained much of their polymorphism in other species of Polistes with six polymorphic loci for P. fuscatus and P. apachus which had a mean H O of 0.48. No polymorphisms were detected outside of the Polistes genus, however, it is likely that some polymorphisms went undetected due to the small number of individuals screened in the other species (Table 1).