Microsatellite genotyping
To document temporal and spatial nuclear genetic variation in H.
horticola wasps, we genotyped 323 wasps (including 195 females) using
the 14 microsatellite-loci and genotyping conditions developed by
Couchoux et al. (Couchoux, Seppä, & van Nouhuys, 2015b). Couchoux et
al. (2015b) demonstrated that there is no linkage disequilibrium between
each pair of these microsatellite loci. The forward primers were
labelled with either FAM, HEX, or TAMRA fluorescent dye (DNA Technology
A/S), and used in multiplex non-overlapping PCR reactions using Qiagen
Multiplex PCR kit (Cat. #206143, Qiagen, USA) as described by Couchoux
et al. (2015b). Diluted PCR products were genotyped on an automated ABI
3730 DNA Sequencer (Applied BiosystemsTM, USA). The
sizes were called using Genescan-500 ROX size standard. We manually
curated the genotypes for each sample using the GeneMapper® Software 5
(Applied BiosystemsTM, USA).
For the purpose of the genetic analyses described below, we separated
the samples into the five localities they were collected from (North
Finström, North Föglö, Seglinge-Kumlinge, Saltvik or Sottunga, Figure
1b), or into 12 spatio-temporal groups according to their geographic
origin and their collection time interval (Finström 1992-97, 2003-08,
2009-11, Föglö 2000-09, 2010-13, Seglinge-Kumlinge 2000-09, 2010-11,
Saltvik 1999, 2005-09, 2010-13, and Sottunga 2002-04, 2005-09; Figure 3,
Table 2). Each of the 12 spatio-temporal groups was designed to include
specimens from roughly either the 90’s, the 2000’s, and the 2010’s, from
each locality, but due to the smaller sample size, the samples from
Sottunga were divided between two temporal groups: the early and the
late 2000’s.