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      Sampling for Microsatellite-Based Population Genetic Studies: 25 to 30 Individuals per Population Is Enough to Accurately Estimate Allele Frequencies

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          Abstract

          One of the most common questions asked before starting a new population genetic study using microsatellite allele frequencies is “how many individuals do I need to sample from each population?” This question has previously been answered by addressing how many individuals are needed to detect all of the alleles present in a population (i.e. rarefaction based analyses). However, we argue that obtaining accurate allele frequencies and accurate estimates of diversity are much more important than detecting all of the alleles, given that very rare alleles (i.e. new mutations) are not very informative for assessing genetic diversity within a population or genetic structure among populations. Here we present a comparison of allele frequencies, expected heterozygosities and genetic distances between real and simulated populations by randomly subsampling 5–100 individuals from four empirical microsatellite genotype datasets ( Formica lugubris, Sciurus vulgaris, Thalassarche melanophris, and Himantopus novaezelandia) to create 100 replicate datasets at each sample size. Despite differences in taxon (two birds, one mammal, one insect), population size, number of loci and polymorphism across loci, the degree of differences between simulated and empirical dataset allele frequencies, expected heterozygosities and pairwise F ST values were almost identical among the four datasets at each sample size. Variability in allele frequency and expected heterozygosity among replicates decreased with increasing sample size, but these decreases were minimal above sample sizes of 25 to 30. Therefore, there appears to be little benefit in sampling more than 25 to 30 individuals per population for population genetic studies based on microsatellite allele frequencies.

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          Estimation of average heterozygosity and genetic distance from a small number of individuals.

          M Nei (1978)
          The magnitudes of the systematic biases involved in sample heterozygosity and sample genetic distances are evaluated, and formulae for obtaining unbiased estimates of average heterozygosity and genetic distance are developed. It is also shown that the number of individuals to be used for estimating average heterozygosity can be very small if a large number of loci are studied and the average heterozygosity is low. The number of individuals to be used for estimating genetic distance can also be very small if the genetic distance is large and the average heterozygosity of the two species compared is low.
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            Do polymorphic loci require large sample sizes to estimate genetic distances?

            The coefficient of variation of estimates of three genetic distances (standard genetic distance of Nei, chord distance, FST) was examined with computer simulation to determine if large samples (per population) are necessary to precisely estimate genetic distances at loci with high levels of polymorphism. These simulations showed that loci with high mutation rates produce estimates of genetic distance with lower coefficients of variation than loci with lower mutation rates--without requiring larger sample sizes from each population. In addition, the rate at which increasing sample sizes decreases the coefficient of variation of estimates of genetic distances was shown to be approximately determined by the value of FST between the populations being sampled. When FST was greater than 0.05, sampling fewer than 20 individuals (per population) should be sufficient. When FST was less than 0.01, sampling 100 individuals (per population) or more will be useful.
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              Impact of landscape management on the genetic structure of red squirrel populations.

              Landscape management practices that alter the degree of habitat fragmentation can significantly affect the genetic structure of animal populations. British red squirrels use "stepping stone" patches of habitat to move considerable distances through a fragmented habitat. Over the past few decades, the planting of a large conifer forest has connected groups of forest fragments in the north of England with those in southern Scotland. This "defragmentation" of the landscape has resulted in substantial genetic mixing of Scottish and Cumbrian genes in squirrel populations up to 100 kilometers from the site of the new forest. These results have implications for the conservation management of animal and plant species in fragmented landscapes such as those found in Britain.
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                Author and article information

                Contributors
                Role: Editor
                Journal
                PLoS One
                PLoS ONE
                plos
                plosone
                PLoS ONE
                Public Library of Science (San Francisco, USA )
                1932-6203
                2012
                12 September 2012
                : 7
                : 9
                : e45170
                Affiliations
                [1 ]School of Biological Sciences, University of Canterbury, Christchurch, New Zealand
                [2 ]Department of Biology, University of Lethbridge, Lethbridge, Canada
                British Columbia Centre for Excellence in HIV/AIDS, Canada
                Author notes

                Competing Interests: The authors have declared that no competing interests exist.

                Conceived and designed the experiments: MLH TMB TES. Analyzed the data: MLH. Wrote the paper: MLH TMB TES. Collected the data: MLH TMB TES.

                Article
                PONE-D-12-11353
                10.1371/journal.pone.0045170
                3440332
                22984627
                345a0a02-3c32-45ea-95b5-eed382667183
                Copyright @ 2012

                This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

                History
                : 19 April 2012
                : 17 August 2012
                Page count
                Pages: 10
                Funding
                The unpublished ant and dataset was collected by MLH and Dr. Kirsten Wolff, funded by the University of Newcastle (UK). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
                Categories
                Research Article
                Biology
                Computational Biology
                Population Genetics
                Genetic Polymorphism
                Ecology
                Conservation Science
                Evolutionary Biology
                Population Genetics
                Genetic Polymorphism
                Evolutionary Genetics
                Genetics
                Heredity
                Genotypes
                Population Genetics
                Gene Pool
                Genetic Polymorphism
                Population Biology
                Population Genetics
                Genetic Polymorphism

                Uncategorized
                Uncategorized

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