Computational protocol: Population genetics of the understory fishtail palm Chamaedorea ernesti-augusti in Belize: high genetic connectivity with local differentiation

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Protocol publication

[…] QIAGEN DNEasy Mini kits (Valencia, CA) were used to isolate DNA from leaf tissue. We characterized nine nuclear microsatellites to measure genetic variation following Cibrián-Jaramillo et al []. Dataset editing and formatting was done with the Excel Microsatellite Toolkit[]. Linkage disequilibrium (LD) [] and deviations from Hardy-Weinberg equilibrium (HWE) could indicate the presence of population structure or inbreeding [], therefore the presence of LD was investigated at the 5% statistical significance level among loci per population based on sampling sites with 10,000 permutations. Departure from HWE expectations was tested for each locus with default parameters; both analyses were carried out in Arlequin[]. Heterozygosity was measured as the unbiased expected heterozygosity (He), and observed heterozygosity (Ho) corrected for sample size []. All genetic variation estimates were obtained in Genetix v4.05 []. The presence of null alleles and allelic dropout was tested using Micro-checker v2.2.1 [], which calculates the probabilities for the observed number of homozygotes within homozygote classes using a cumulative binomial distribution []. [...] We tested for recent subdivision using BayesAss+ v1.3 []. This method estimates a matrix of pairwise recent immigration rates among populations, m, using a coalescent approach. This approach assumes that the estimated clusters are a result of gene flow in very recent generations []. Default settings for burn-in and number of MCMC iterations were enough to reach convergence based on visual inspection of likelihood scores. Five independent runs were performed to test for congruence, and a likelihood ratio test was then employed to determine whether the prior and posterior probabilities of migration rates are significantly different from each other []. The presence of linkage among some loci and the overlapping generations of C. ernesti-augusti could decrease the accuracy of each estimate, but not necessarily change the relative proportions of m among localities.Small and contrasting sample sizes may confound estimates in this method, so we compared BayesAss+ with individual assignments using the frequency criterion of Paetkau et al [], implemented in GeneClass v2.0 []. The frequency method approximates the distribution of genotype likelihoods of the individual that will be assigned to the simulated distribution of a reference population. It takes into account type I error and reduces the amount of resident individuals being excluded, as well as incorporating the sampling variance that results from different population sizes []. [...] Limited dispersal can result in a pattern of genetic differentiation that increases with geographic distance. Isolation by distance (IBD) was tested with a Mantel test for matrix correspondence [,]. Genalex v.6 was used to produce a geographic and genetic distance matrix using pairwise individual comparisons, following [] and []. A Mantel test with populations using [] linearized distance Fst/(1-Fst) was also calculated on the IBD webserver available at []. […]

Pipeline specifications

Software tools Arlequin, GeneClass, GenAlEx
Application Population genetic analysis
Organisms Homo sapiens