Automated genotyping of microsatellite loci from feces with high throughput sequences
Autor: | Jennifer A. Leonard, Isabel Salado, Carles Vilà, Alberto Fernández-Gil |
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Přispěvatelé: | Ministerio de Ciencia, Innovación y Universidades (España) |
Jazyk: | angličtina |
Rok vydání: | 2021 |
Předmět: |
Heredity
Artificial Gene Amplification and Extension Polymerase Chain Reaction Homozygosity law.invention Feces Sequencing techniques law Genotype Multiplex DNA sequencing Throughput (business) Polymerase chain reaction education.field_of_study Multidisciplinary Heterozygosity Software Engineering High-Throughput Nucleotide Sequencing Genomics Genetic Mapping Microsatellite Engineering and Technology Medicine Transcriptome Analysis Research Article Next-Generation Sequencing Genotyping Computer and Information Sciences Science Population Variant Genotypes Computational biology Biology Research and Analysis Methods Computer Software Genetics Animals education Molecular Biology Techniques Molecular Biology Wolves Software Tools Biology and Life Sciences Computational Biology Genome Analysis Genetic Loci Microsatellite Repeats |
Zdroj: | PLoS ONE, Vol 16, Iss 10, p e0258906 (2021) PLoS ONE PLoS ONE, Vol 16, Iss 10 (2021) Digital.CSIC. Repositorio Institucional del CSIC instname |
Popis: | Ecological and conservation genetic studies often use noninvasive sampling, especially with elusive or endangered species. Because microsatellites are generally short in length, they can be amplified from low quality samples such as feces. Microsatellites are highly polymorphic so few markers are enough for reliable individual identification, kinship determination, or population characterization. However, the genotyping process from feces is expensive and time consuming. Given next-generation sequencing (NGS) and recent software developments, automated microsatellite genotyping from NGS data may now be possible. These software packages infer the genotypes directly from sequence reads, increasing throughput. Here we evaluate the performance of four software packages to genotype microsatellite loci from Iberian wolf (Canis lupus) feces using NGS. We initially combined 46 markers in a single multiplex reaction for the first time, of which 19 were included in the final analyses. Megasat was the software that provided genotypes with fewer errors. Coverage over 100X provided little additional information, but a relatively high number of PCR replicates were necessary to obtain a high quality genotype from highly unoptimized, multiplexed reactions (10 replicates for 18 of the 19 loci analyzed here). This could be reduced through optimization. The use of new bioinformatic tools and next-generation sequencing data to genotype these highly informative markers may increase throughput at a reasonable cost and with a smaller amount of laboratory work. Thus, high throughput sequencing approaches could facilitate the use of microsatellites with fecal DNA to address ecological and conservation questions |
Databáze: | OpenAIRE |
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