Computational protocol: Taxonomic status and molecular phylogeography of two sibling species of Polytremis (Lepidoptera: Hesperiidae)

Similar protocols

Protocol publication

[…] The sequence data of the mitochondrial COI and nuclear wingless were aligned with published homologous sequence from genus Polytremis (e. g., P. eltola Hewitson, 1869, P. discreta Elwes & Edwards, 1897, etc.) as well as Borbo cinnara Wallace, 1866 and Pseudoborbo bevani Moore, 1878 (presented in ) respectively, translated to amino acid sequences to check for nuclear mitochondrial pseudogenes (numts) and pruned to remove redundant sequences with Bioedit v.7.0. The haplotype sequence matrix was used for all subsequent phylogenetic analyses (). Phylogenetic trees were constructed by the ML (maximum-likelihood) methods with PhyML. Modeltest 3.7 was used to select the optimal nucleotide substitution models following the Akaike Information Criterion (AIC). In the ML analysis, a heuristic search was conducted. The starting tree for branch-swapping was from stepwise addition. Nodal support of the ML tree was estimated by 1000 bootstraps. Network profile of the haplotypes identified in P. t. theca, P. t. fukia, P. nascens and P. mencia was constructed with Network4.5 using the median-joining method. The haplotype diversity (Hd) and nucleotide diversity (π) for P. t. theca and P. t. fukia were estimated by DnaSP4.90.For the COI data set, pairwise FST was also calculated with Arlequin v3.0, which accurately reflects patterns of genetic variation. Correspondingly, gene flow was estimated in Arlequin v3.0. Two level hierarchical analyses of molecular variance (AMOVA) were conducted to evaluate possible population genetic structure of P. t. theca and P. t. fukia using Arlequin v3.0 with 1,000 permutations. We calculated Tajima’s D and Fu’s F statistic and ran 10,000 coalescent simulations for each statistic to create 95% confidence intervals investigate the historical population demographics and testing whether the sequences conformed to the expectations of neutrality. Pairwise mismatch distribution analyses were performed for all P. theca specimens and two subspeices specimens separately to find the evidence of past demographic expansions using DnaSP4.90. The times to the most recent common ancestor of the major lineages and the whole population were estimated using relax-clock molecular dating estimation implemented in the BEAST 1.5.2. Analyses using the HKY model of nucleotide substitution with gamma distributed rate variation among sites were performed. The Yule speciation method was assumed and the nucleotide substitution rate of 3.54% per million years that has generally been calibrated for COI in insect was used. Chains were run for 50 million generations, with the first 20% discarded as burn-in. The results were summarized through TRACER 1.5. […]

Pipeline specifications