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Each tree was rooted using ML by selecting the rooted tree with the best likelihood under the molecular clock constraint or by outgroup rooting using strains from the other subject.
Trees were rooted by ML rooting by selecting the rooted tree with the best likelihood under the molecular clock constraint, or by outgroup rooting using the earliest PBMC samples as outgroup.
Trees were rooted by ML rooting by selecting the rooted tree with the best likelihood under the molecular clock constraint (taking into account the different sampling times of the taxa) [30], [31], or by outgroup rooting using two Albanian HIV-1A strains as outgroup for the HIV-1B data set, and two HIV-1B Albanian strains for the HIV-1A data sets.
FigTree v.1.3.1 [ 52] was used to display summarized and annotated phylogenetic trees with a molecular clock constraint yielded by BEAST.
They analysed these datasets taking a Bayesian relaxed molecular clock approach combined with fossil calibrations and global molecular clock constraint conditions.
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To evaluate whether the Ligia sequences examined exhibit clocklike behavior, we used a Likelihood Ratio Test on the likelihood scores obtained with PAUP* [v. 4b10, 41] of the tree in Figure 3 with and without molecular clock constraints.
To estimate the divergence time among the AGOs, the molecular clock test was performed by comparing the ML values for a given tree topology with molecular clock constraints to a topology, and without the molecular clock constraints under the JTT (+G + I) model [ 50] by using MEGA 5.2.
The molecular clock hypothesis was not violated, as likelihood scores of trees constructed with and without molecular clock constraints were not significantly different (χ2 = 44.8 d.f. = 176, P > 0.05).
We tested whether our Acacia consensus tree obeyed a molecular clock hypothesis using MEGA5 by comparing the ML value for our topologies with and without the molecular clock constraints using the GTR+I+Γ model of evolution.
In order to test if the mtDNAs evolved clock like a likelihood ratio test was performed in MEGA5 (Tamura et al., 2011) by comparing the maximum likelihood (ML) value for the given topology using only the modern strains with and without the molecular clock constraints.
For the calibration of the molecular clock, stringent constraints were applied based on fossils that can be unambiguously assigned to extant clades and whose unique skeletal characters can be unequivocally recognized in fossil coralla.
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