Application of a SSR‐GBS marker system on investigation of European Hedgehog species and their hybrid zone dynamics

Abstract

By applying second‐generation sequencing technologies to microsatellite genotyping, sequence information is produced which can result in high‐resolution population genetics analysis populations and increased replicability between runs and laboratories. In the present study, we establish an approach to study the genetic structure patterns of two European hedgehog species Erinaceaus europaeus and E. roumanicus . These species are usually associated with human settlements and are good models to study anthropogenic impacts on the genetic diversity of wild populations. The short sequence repeats genotyping by sequence (SSR‐GBS) method presented uses amplicon sequences to determine genotypes for which allelic variants can be defined according to both length and single nucleotide polymorphisms (SNPs). To evaluate whether complete sequence information improved genetic structure definition, we compared this information with datasets based solely on length information. We identified a total of 42 markers which were successfully amplified in both species. Overall, genotyping based on complete sequence information resulted in a higher number of alleles, as well as greater genetic diversity and differentiation between species. Additionally, the structure patterns were slightly clearer with a division between both species and some potential hybrids. There was some degree of genetic structure within species, although only in E. roumanicus was this related to geographical distance. The statistically significant results obtained by SSR‐GBS demonstrate that it is superior to electrophoresis‐based methods for SSR genotyping. Moreover, the greater reproducibility and throughput with lower effort which can be obtained with SSR‐GBS and the possibility to include degraded DNA into the analysis, allow for continued relevance of SSR markers during the genomic era.

Molecular genetic diversity and differentiation of Nile tilapia (Oreochromis niloticus, L. 1758) in East African natural and stocked populations

“…Genomic DNA extraction was conducted using magnetic beads based on the MagSi-DNA Vegetal kit (MagnaMedics, Geleen, Netherlands) and a magnetic separator, SL-MagSep96 (Steinbrenner, Germany) [27,30]. We used microsatellite markers [27], to which we added 15 extra primers ( Table 2, see also Additional file 1: Table S3).…”

“…The raw sequence data were deposited in the GenBank, sequence read archive database (SRA) under the project PRJNA550300 with the accession numbers, SRR9587388 to SRR9587270. Sequences generated by Illumina, were subsequently quality checked and controlled, which were later used for alleles calling as described in [27,30] using the scripts from the SSR-GBS pipeline (https://github.com/mcurto/SSR-GBS-pipeline). The resulting codominant matrix and information for which sequences correspond to each allele can be found in the Additional file 2 (see the file named "Second_additional fileAllelesList & matrix_").For subsequent analyses, all loci and samples with missing genotypes ≥50% were excluded, leaving a total number of 40 markers (Additional file 1: Tables S1, S3).…”

“…SSR-GBS approaches are useful because they reduce size homoplasy, which is one of the constraints of traditional SSR fragment length analysis [28,29]. However, SSR-GBS is not without drawbacks [30]. For example, the presence of stutter complicates allele calling for dinucleotides, null alleles due to mutation on primer binding sites, and it does not recover genomic information hence overestimating events that had a small impact on the gene-pool.…”

Pollen availability for the Horned mason bee (Osmia cornuta) in regions of different land use and landscape structures

“…On Sanger sequence‐based DNA barcoding, longer regions are collected (up to 800 bp), and consequently, these were chosen for pollen barcoding (CBOL Plant Working Group, ). Because the length for Illumina amplicon sequencing cannot exceed 500 bp (Curto et al ., ), mostly the chloroplast region rbcl and the ribosomal DNA marker ITS are preferably used (e.g. Richardson et al ., ; Bell et al ., 2017; Suchan et al ., ).…”

“…Paired reads were merged using pear (Zhang et al ., ) and amplification primers were trimmed using script 1 from Curto et al . (). The resulting reads were dereplicated and blast ed against all Spermatophyta chloroplast sequences present in GenBank.…”