Additional file 1:

Maximum likelihood phylogeny based on 4813 bp of the combined Mll, Myh6, Ptr, Rag1, Rag2, and Ryr3 genes. Numbers above branches indicate maximum parsimony and maximum likelihood bootstrap values, followed by Bayesian posterior probabilities.

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Additional file 2:

Vertebral count for each taxa identified on the Cyt b phylogeny.

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Additional file 3:

Estimated ages of the most recent common ancestor of interstitial Luciogobius species. Every node on the ultrametric tree obtained by penalized likelihood was used individually as a calibration point to obtain a conservative range of estimates for the most recent common ancestor of the interstitial Luciogobius species. Optimal ages and 95% credibility intervals are plotted against the relative node heights of those used to calibrate the chronogram. The 7.67 Ma estimate obtained based on Gymnogobius isaza-G. urotaenia split (2.3 Ma) is given by the dotted line, and 95% credibility interval of this estimate is indicated by the shade.

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Additional file 4:

Optimal age of the root node inferred based on Cyt b substitution rate. The age of the root node (Eutaeniichthys gilli-Typhlogobius californiensis split) was adjusted using an iterative approach until average substitution rates across branches equaled the known rate of 0.0135 substitutions/site/My [24].

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Additional file 5:

Median gravel size and sorting index for five interstitial Luciogobius species.

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Additional file 6:

List of species examined and locality information.

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Additional file 7:

List of primers used in this study.

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Additional file 8:

Plots of uncorrected pairwise transition distances against transversion distances in each of the three Cyt b codon positions.

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