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The bootstrap consensus tree inferred from 500 replicates was constructed to represent the evolutionary history of the HKT genes by using the Minimum Evolution Method in MEGA5 (Tamura et al. 2011).
The phylogenetic tree was produced by the minimum evolution method and 1000 bootstrap iteration.
Phylogenetic trees based on multiple alignments of nucleotide sequences were constructed using the minimum evolution method as implemented in the MEGA program with default parameters.
Phylograms were constructed by Mega4.1 using the Minimum Evolution method with a bootstrap test based on 5000 replicates, and the Poisson correction method [5].
The phylogram represents a consensus tree based on 100 bootstrap iterations, calculated by the Minimum Evolution method under default parameters [66].
In case of FAFLP data, the trees were constructed using the binary data by minimum evolution method with topology validated through an interior branch test, a t-test, which is computed using the bootstrap procedure.
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Instead, the strains branched deeply within the Proteobacteria as shown in a phylogenetic tree created by maximum likelihood [19], Figure 3. Very similar tree topologies were found by neighbor-joining, maximum parsimony or minimum evolution methods using MEGA3 [20](data not shown).
The neighbour-joining and minimum evolution methods were then used to plot the phylogenetic trees (pair-wise deletion, JonesTaylor-Thornton matrix and enter range activated sites (gamma-number 2.5).
Phylogenetic relationships were determined by using the Jukes-Cantor algorithm combined with either neighbor-joining (NJ) or Minimum Evolution methods implemented in MEGA.
To further control the reliability of the maximum likelihood approach, the phylogenetic tree calculations were repeated using the neighbor joining and minimum evolution methods implemented in MEGA6.
Neighbor-joining and minimum evolution methods in MEGA4 (http://www.megasoftware.net) were used to construct the trees along with multiple substitution models, including p-distance and Jukes-Cantor; all methods gave similar results.
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