Computational protocol: Phylogeography of the Chinese Beard Eel, Cirrhimuraena chinensis Kaup, Inferred from Mitochondrial DNA: A Range Expansion after the Last Glacial Maximum

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Protocol publication

[…] cyt b and CR sequences were edited and aligned in Sequencher version 4.1.4 (Gene Codes Corporation, Ann Arbor, MI, USA) before being incorporated into the concatenated dataset. Genetic diversity estimates of cyt b, CR sequences and the concatenated dataset, including the haplotype diversity (h) and nucleotide diversity (π), were calculated for each location and the entire population by DnaSP version 5.0 [].To evaluate the genetic differentiation between sampling locations, population genetic distances (FST) were estimated using concatenated data by Arlequin version 3.5 []. Analysis of molecular variation (AMOVA) was implemented in Arlequin to investigate the partition of genetic variation within and among sampling locations, along with hierarchical analysis by population groupings to investigate the possible effects of major potential geographical barriers located in the sampling range, including the Taiwan Strait, the Pearl River, Leizhou Peninsula and the Qiongzhou Strait, on the phylogeographic pattern: (1) Ningde as a single group (ND), with the remaining populations on the other side of the Taiwan Strait constituting another group; (2) populations were divided into an eastern group (ND, XM, ST) and a western group (YJ, BH, HK, SY) at the boundary of the Pearl River; (3) Leizhou Peninsula spilt the populations into an eastern group (ND, XM, ST, YJ) and a western group (BH, HK, SY); (4) populations on Hainan island belonged to an island group (HK, SY), while the mainland group (ND, XM, ST, YJ) contained the remainder in the case that the Qiongzhou Strait served as a vicariant barrier. A Mantel test was conducted to assess the significance of the relationship between the pairwise FST matrix and the matrix of geographic distance.Historical demography was inferred using three approaches. First, DnaSP was used to conduct neutrality tests of Tajima’s D [], Fu’s FS [] and Ramos–Onsins and Rozas’ R2 [] to detect whether there was any deviation from the assumption of neutrality, which would indicate a recent population expansion. Tajima’s D is widely used in neutrality tests, and Fu’s FS and the Ramos–Onsins and Rozas R2 test were confirmed to be the most powerful tools in examining population growth []. The latter was proven to be particularly sensitive for limited sample sizes []. Second, population demographic dynamics were investigated through nucleotide mismatch distribution analysis [,,,] by DnaSP. In addition, the sum of square deviation (SSD) and Harpending’s raggedness index were also calculated by Arlequin. We also calculated the time since population expansion using the equation, t = τ/2μ, where μ represents the mutation rate of the marker used and τ is a parameter obtained from mismatch distribution analysis in Arlequin. Finally, Bayesian skyline plots were created with BEAST version 1.7.4 []. As the mutation rate of D-loop ranged from a minimum 2.2% per million years (myr) in Cichlidae [] to a maximum of 20% in Clupeidae [], while that of cyt b ranged from 0.68%–2% per myr [,], we took 5% per myr [] as the putative mutation rate for the concatenated sequences. We employed a GTR model with parameters for invariable sites and gamma distribution (GTR + I + G) and ran 106 generations, discarding 105 generations as burn-in. The result was subsequently visualized by Tracer 1.5 []. […]

Pipeline specifications

Software tools Sequencher, DnaSP, Arlequin, BEAST
Application Population genetic analysis
Organisms Eucryptorrhynchus scrobiculatus