Fast and cost‐effective single nucleotide polymorphism (<scp>SNP</scp>) detection in the absence of a reference genome using semideep next‐generation Random Amplicon Sequencing (<scp>RAM</scp>seq)

<jats:title>Abstract</jats:title><jats:p>Biodiversity has suffered a dramatic global decline during the past decades, and monitoring tools are urgently needed providing data for the development and evaluation of conservation efforts both on a species and on a genetic level. However, in wild species, the assessment of genetic diversity is often hampered by the lack of suitable genetic markers. In this article, we present Random Amplicon Sequencing (<jats:styled-content style="fixed-case">RAM</jats:styled-content>seq), a novel approach for fast and cost‐effective detection of single nucleotide polymorphisms (<jats:styled-content style="fixed-case">SNP</jats:styled-content>s) in nonmodel species by semideep sequencing of random amplicons. By applying <jats:styled-content style="fixed-case">RAM</jats:styled-content>seq to the Eurasian otter (<jats:italic>Lutra lutra</jats:italic>), we identified 238 putative <jats:styled-content style="fixed-case">SNP</jats:styled-content>s after quality filtering of all candidate loci and were able to validate 32 of 77 loci tested. In a second step, we evaluated the genotyping performance of these <jats:styled-content style="fixed-case">SNP</jats:styled-content> loci in noninvasive samples, one of the most challenging genotyping applications, by comparing it with genotyping results of the same faecal samples at microsatellite markers. We compared (i) polymerase chain reaction (<jats:styled-content style="fixed-case">PCR</jats:styled-content>) success rate, (ii) genotyping errors and (iii) Mendelian inheritance (population parameters). <jats:styled-content style="fixed-case">SNP</jats:styled-content>s produced a significantly higher <jats:styled-content style="fixed-case">PCR</jats:styled-content> success rate (75.5% vs. 65.1%) and lower mean allelic error rate (8.8% vs. 13.3%) than microsatellites, but showed a higher allelic dropout rate (29.7% vs. 19.8%). Genotyping results showed no deviations from Mendelian inheritance in any of the <jats:styled-content style="fixed-case">SNP</jats:styled-content> loci. Hence, <jats:styled-content style="fixed-case">RAM</jats:styled-content>seq appears to be a valuable tool for the detection of genetic markers in nonmodel species, which is a common challenge in conservation genetic studies.</jats:p>