1

Chapter 2: Amino Acid Substitutions in Euteleost

Rhodopsins Correlate with Habitat Depth: Linking

Molecular Changes to Ecology of Organisms

Table S1. List of sites in rhodopsin that correlated with habitat depth, using codon or unity

branch lengths, in at least one method. Sites which were identified using the time-calibrated

phylogeny are shown in bold. Significance was determined by comparing p-values for each site

with the distribution obtained from simulations: *: p < 0.05, **: p < 0.01, ***: p < 0.001, PGLS:

Phylogenetic Generalized Least Squares, DISC: Discrete, AR: Ancestral Reconstructions, KW:

Kruskal-Wallis

Site PGLS-codon DISC-codon AR-codon PGLS-1 DISC-1 AR-1

38 * *

46 ** *

50 *

63 **

83 * **

88 *

95 *

97 ** ** ** ***

112 *

114 ** ** *

119 **

122 *

124 * *

130 *

137 ** *

144 *** * *

149 *

159 *

160 ** *

166 *** * * *

168 ** *

189 * *

196 *

209 ***

210 *

214 *

217 ** ** **

219 *

251 ** ***

259 *

266 ** ** ***

270 *

2

271 * *

273 *

274 *

277 **

278 **

281 ** *

282 *

292 *** *** *** *** *** ***

298 *

300 *

Table S2. Number and proportion (in brackets) of sites with significant (p < 0.05) correlation

with habitat depth in full and pruned rhodopsin data sets. Number of sequences for each pruned

data set is shown followed by the name of the gene, in square brackets ([]), to which the pruned

RH1 data set corresponds. Significance was determined by comparison with simulated

distributions. PGLS: Phylogenetic Generalized Least Squares, DISC: Discrete, AR: Ancestral

Reconstructions, KW: Kruskal-Wallis

Num. of species PGLS DISC AR KW

196 [RH1] 15 (17.6% ) 7 (8.2%) 16 (18.8%) 3 (3.5%)

173 [COI] 12 (14.3%) 7 (8.3%) 14 (16.7%) 2 (2.4%)

134 [CYTB] 7 (9.5%) 3 (4.1%) 8 (10.8%) 1 (1.4%)

115 [RAG1] 13 (17.1%) 2 (2.6%) 8 (10.5%) 0 (0%)

110 [MYH6] 15 (19.7%) 5 (6.6%) 9 (11.8%) 1 (1.3%)

105 [ZIC1] 7 (10.0%) 2 (2.9%) 8 (11.4%) 1 (1.4%)

100 [MLL] 7 (10.9%) 2 (3.1%) 5 (7.8%) 1 (1.6%)

87 [ND2] 7 (10.9%) 1 (1.6%) 9 (14.1%) 0 (0%)

74 [ND1] 8 (14.5%) 1 (1.8%) 10 (18.2%) 0 (0%)

73 [SH3PX3] 7 (11.1%) 3 (4.8%) 6 (9.5%) 0 (0%)

67 [ENC1] 8 (13.3%) 1 (1.7%) 5 (8.3%) 0 (0%)

56 [ND4] 6 (11.5%) 1 (1.9%) 14 (26.9%) 0 (0%)

54 [RAG2] 3 (6.4%) 0 (0%) 5 (10.6%) 0 (0%)

53 [ND5] 4 (8.2%) 1 (2.0%) 10 (20.4%) 0 (0%)

Table S3. Comparison of five models tested in this study to describe the evolution of habitat

depths in euteleosts, using the time-calibrated phylogeny. Ancestral depths were reconstructed

under Brownian Motion, Ornstein-Uhlenbeck, and Pagel’s Lambda (lowest AICc) models.

BM OU Lambda Kappa Delta

ln L -1390.9 -1387.4 -1386.1 -1388.0 -1388.0

AICc 2785.9 2781.0 2778.4 2782.1 2782.1

Parameter α = 0.005 λ = 0.913 κ = 0.725 δ = 2.572

3

Table S4. Comparison of false positive rate (type I error) of each approach, as determined by

simulations, across different branch lengths at nominal α = 0.05. PGLS: Phylogenetic

Generalized Least Squares, DISC: Discrete, AR: Ancestral Reconstructions, KW: Kruskal-

Wallis

Non-phy KW PGLS DISC AR

Time BL 63.7% 8.7% 22.1% 0.6%

Codon BL 63.7% 3.9% 13.0% 0.6%

Unity BL 63.7% 16.0% 12.4% 0.7%

Table S5. Comparison of number and proportion (in brackets) of sites in rhodopsin with

significant (p < 0.05) correlation with habitat depth across different branch lengths. Non-phy

KW: Kruskal-Wallis test without using phylogeny, PGLS: Phylogenetic Generalized Least

Squares, DISC: Discrete, AR: Ancestral Reconstructions, KW: Kruskal-Wallis test after

comparison with a simulated distribution

Non-phy KW PGLS DISC AR KW

Time BL 55 (64.7%) 15 (17.6% ) 7 (8.2%) 16 (18.8%) 3 (3.5%)

Codon BL 55 (64.7%) 16 (18.8%) 13 (15.3%) 12 (14.1%) 3 (3.5%)

Unity BL 55 (64.7%) 6 (7.1%) 11 (12.9%) 11 (12.9%) 3 (3.5%)

4

Table S6. Accession numbers of sequences downloaded from NCBI.

Species RH1 COI CYT

B

MY

H6

ML

L

ND1 ND2 ND4 ND5 RAG

1

RAG

2

SH3

PX3

ZIC1 ENC

1

Acanthocybi

um_solandri

DQ87480

4.1

JQ840406.

1

DQ197922.

1

DQ854691.

1

DQ87477

2.1

KC83109

3.1

Aeoliscus_str

igatus

EU637931.

1

NC_010270

.1

NC_010270

.1

JX189647.

1

NC_010270

.1

NC_010270

.1

NC_010270

.1

NC_010270

.1

JX189801.

1

JX189554.

1

JX189030.

1

JX188862.

1

Aethotaxis_

mitopteryx

DQ49878

1.1

JN640756.

1

JF264480.1

JN187038.

1

DQ49876

6.1

DQ367401.

1

JN186956.

1

JN186820.

1

Alepocephalu

s_bairdii

JN412584.

1

NC_013567

.1

NC_013567

.1

NC_013567

.1

NC_013567

.1

NC_013567

.1

NC_013567

.1

Amblyeleotri

s_guttata

HQ5

3691

2.1

FJ582

710.1

KC8

2692

4.1

HQ53

6711.

1

FJ796

083.1

HQ5

3681

6.1

KC8

3076

7.1

Amblyeleotri

s_wheeleri

HQ5

3691

8.1

HQ56

1506.

1

KC8

2692

5.1

AF39

1455.

1

AF39

1527.

1

HQ5

3686

5.1

Amblygobius

_phalaena

HQ5

3689

7.1

JQ43

1409.

1

KF41

5498.

1

KC8

2693

6.1

AF39

1450.

1

AF39

1522.

1

KF41

5693.

1

HQ5

3680

3.1

KF41

5897.

1

Ammodytes_

tobianus

AY1

4130

6.1

AY3

6223

4.1

Amphiprion_

melanopus

HM1

0782

4.1

KF92

9596.

1

DQ34

3955.

1

EU2

5700

8.1

Anarhichas_l

upus

EU63

7936.

1

NC_0

09773

.1

NC_0

09773

.1

JX18

9742.

1

EU6

3802

6.1

NC_0

09773

.1

NC_0

09773

.1

NC_0

09773

.1

NC_0

09773

.1

JX18

9900.

1

JX18

9494.

1

JX18

9127.

1

JX18

8973.

1

Anoplogaster

_cornuta

JN41

2582.

1

NC_0

04391

.1

NC_0

04391

.1

KF13

9922.

1

NC_0

04391

.1

NC_0

04391

.1

NC_0

04391

.1

NC_0

04391

.1

KF14

1170.

1

KF14

1400.

1

KF14

0476.

1

KF13

9397.

1

Antigonia_ca

pros

EU63

7937.

1

NC_0

03191

.1

NC_0

03191

.1

KF02

7821.

1

EU6

3802

7.1

NC_0

03191

.1

NC_0

03191

.1

NC_0

03191

.1

NC_0

03191

.1

AY3

0878

5.1

KF02

8051.

1

EF53

3930.

1

EF53

9254.

1

Aphanopus_c

arbo

EU63

7938.

1

NC_0

22506

.1

NC_0

22506

.1

EU6

3802

8.1

NC_0

22506

.1

NC_0

22506

.1

NC_0

22506

.1

NC_0

22506

.1

KC8

3111

1.1

Apogon_qua

drifasciatus

EU63

7940.

1

EU14

8589.

1

Apsilus_fusc

us

EU63

7941.

1

EU6

3802

9.1

Argyropelecu

s_gigas

JN41

2572.

1

GQ86

0365.

1

GQ8

6047

4.1

GQ8

6031

2.1

JX19

1296.

1

GQ8

6042

1.1

Arnoglossus_

imperialis

AY1

4128

3.1

EU51

3606.

1

EU51

3758.

1

JQ93

9581.

1

AY3

6222

8.1

JQ93

8321.

1

JQ94

0213.

1

Asterroptery

x_ensifera

HQ5

3693

1.1

JQ34

9787.

1

HQ90

9506.

1

FJ796

099.1

HQ5

3683

7.1

Aulostomus_

chinensis

AY1

4127

9.1

NC_0

10269

.1

NC_0

10269

.1

KF13

9935.

1

AY3

6222

6.1

NC_0

10269

.1

NC_0

10269

.1

NC_0

10269

.1

NC_0

10269

.1

KF14

1181.

1

KF14

1413.

1

KF14

0488.

1

Auxis_thazar

d

DQ8

7480

1.1

AB10

5447.

1

AB10

5447.

1

AB10

5447.

1

AB10

5447.

1

AB10

5447.

1

AB10

5447.

1

DQ8

7476

9.1

Balistes_capr

iscus

DQ8

74818.1

KC50

0312.1

DQ19

7928.1

KF02

5805.1

HQ59

9505.1

AY7

00308.1

DQ8

74786.1

KF14

1417.1

KF14

0493.1

KF13

9639.1

Balistes_vetu

la

KF02

7975.

AP00

9204.

AP00

9204.

KF13

9941.

KF0

2778

AP00

9204.

AP00

9204.

AP00

9204.

AP00

9204.

AY7

0031

KF02

8057.

KF14

0494.

KF13

9638.

5

1 1 1 1 8.1 1 1 1 1 0.1 1 1 1

Barbourisia_

rufa

AY36833

3.1

NC_012046

.1

NC_012046

.1

JX190468.

1

AY36226

4.1

NC_012046

.1

NC_012046

.1

NC_012046

.1

NC_012046

.1

JX190855.

1

JX190989.

1

JX191314.

1

KF139388.

1

Bathygobius_

curacao

HQ53689

2.1

HM748428

.1

JN575297.

1

AF391426.

1

AF391498.

1

JN575286.

1

HQ53679

6.1

Bathylagus_e

uryops

AY14125

5.1

KF929651.

1

JX190424.

1

JN230895.

1

JX190953.

1

JX191276.

1

Bathypterois

_dubius

AY14125

7.1

KC015247.

1

AY36221

9.1

Benthosema_

suborbitale

JN412576.

1

Beryx_splend

ens

AY1

4126

5.1

NC_0

03188

.1

NC_0

03188

.1

AY3

6223

8.1

NC_0

03188

.1

NC_0

03188

.1

NC_0

03188

.1

NC_0

03188

.1

EF09

5636.

1

Bothus_poda

s

AY3

6831

3.1

EU51

3621.

1

EU51

3764.

1

EU6

3803

3.1

Bovichtus_va

riegatus

AY1

4129

9.1

DQ52

6445.

1

JN18

7019.

1

AY3

6228

3.1

JN18

6882.

1

JN18

6937.

1

JN18

6801.

1

Caesio_diagr

amma

EU63

8000.

1

KF00

9652.

1

EU6

3808

5.1

Callionymus

_lyra

AY1

4127

0.1

JQ77

4796.

1

AY3

6222

5.1

Capros_aper AY1

4126

2.1

NC_0

10958

.1

NC_0

10958

.1

KF13

9953.

1

AY3

6223

3.1

NC_0

10958

.1

NC_0

10958

.1

NC_0

10958

.1

NC_0

10958

.1

EF09

5638.

1

KF14

0506.

1

KF13

9623.

1

Cataetyx_lati

ceps

EU63

7947.

1

KC01

5261.

1

EU6

3803

5.1

Centropomus

_viridis

JQ93

8019.

1

DQ31

0724.

1

AF01

8603.

1

JQ93

9570.

1

JQ93

9459.

1

EU16

7816.

1

JQ94

0179.

1

Cepola_macr

ophthalma

EU63

7948.

1

JQ77

4997.

1

EU6

3803

7.1

EU16

7817.

1

Ceratias_hol

boelli

AY1

4126

3.1

GU44

0269.

1

AY29

2604.

1

KF13

9959.

1

AY3

6227

0.1

KF14

1432.

1

KC8

3081

6.1

Ceratoscopel

us_warmingii

JN41

2573.

1

GU07

1724.

1

KC8

2730

0.1

Cerdale_flori

dana

HQ5

3688

7.1

JQ83

9992.

1

EU38

0922.

1

AF39

1409.

1

AF39

1481.

1

KC8

3076

9.1

Chaetodon_s

emilarvatus

AY3

6831

2.1

AJ74

8305.

1

AY3

6224

0.1

EF09

5655.

1

EF61

7084.

1

Champsodon

_snyderi

EU63

7949.

1

JQ68

1321.

1

KF13

9962.

1

EU6

3803

8.1

KF13

9578.

1

Chauliodus_s

loani

JN41

2563.

1

NC_0

03159

.1

NC_0

03159

.1

KF13

9965.

1

NC_0

03159

.1

NC_0

03159

.1

NC_0

03159

.1

NC_0

03159

.1

GQ8

6032

7.1

KF14

1438.

1

KF14

0514.

1

GQ8

6044

0.1

Cheilopogon

_heterurus

EU63

7950.

1

EU6

3803

9.1

Chionodraco

_hamatus

DQ4

98792.1

HQ91

8335.1

HM0

49940.1

HQ1

69805.1

AY3

62280.1

HQ17

0102.1

HM0

50097.1

HQ1

69986.1

HQ1

70192.1

HQ1

69685.1

HM0

49981.1

Chloroscomb

rus_chrysuru

AY1

4131

JQ84

1500.

AY05

0737.

AY3

6222

KC8

3122

6

s 3.1 1 1 3.1 6.1

Chrysiptera_

cyanea

AB627088.

1

FJ583182.1

AB018992.

1

AB018977.

1

AY20864

3.1

FJ616770.

1

Citharus_lin

guatula

AY14132

3.1

KC500412.

1

EF439511.

1

JQ939634.

1

AY36223

2.1

JQ938324.

1

JQ940218.

1

Coryphaena_

hippurus

DQ87482

4.1

AB355908.

1

AB355908.

1

JX190488.

1

AB355908.

1

AB355908.

1

AB355908.

1

AB355908.

1

KF311976.

1

DQ87479

2.1

JX191010.

1

JX191333.

1

KF139497.

1

Coryphaenoi

des_rupestris

AY36831

9.1

AJ517502.

1

EU001915.

1

EU63804

1.1

FJ215233.

1

EU002077.

1

EU002025.

1

Cottoperca_g

obio

AY14130

0.1

EU074394.

1

JN187021.

1

JN186884.

1

JN186939.

1

JN186803.

1

Cottunculus_

thomsonii

AY3

6831

5.1

KC01

5300.

1

AY3

6226

0.1

Cryptocentru

s_cinctus

HQ5

3692

1.1

FJ583

282.1

KF41

5536.

1

HQ53

6748.

1

FJ796

110.1

KF41

5732.

1

HQ5

3684

9.1

KF41

5940.

1

Ctenochaetus

_striatus

AY1

4132

0.1

JQ34

9928.

1

KC62

3714.

1

EF53

6296.

1

AY3

6224

2.1

EF53

0096.

1

EF53

3919.

1

EF53

9243.

1

Ctenogobiops

_mitodes

HQ5

3694

5.1

HQ53

6671.

1

GU18

7228.

1

GU18

7266.

1

HQ5

3685

3.1

Cubiceps_gr

acilis

EU63

7952.

1

KC01

5307.

1

EU6

3804

2.1

KC8

3103

2.1

Cyclopterus_

lumpus

AY3

6831

6.1

KC01

5314.

1

EU49

2084.

1

JX18

9653.

1

AY3

6221

8.1

JX18

9807.

1

JX18

9560.

1

JX18

9036.

1

JX18

8868.

1

Cynoscion_re

galis

EF09

5618.

1

EU18

0145.

1

GQ22

0003.

1

KF13

9976.

1

EF09

5660.

1

KF14

1449.

1

KF14

0522.

1

KF13

9551.

1

Dactylopteru

s_volitans

AY1

4128

2.1

GU70

2460.

1

EF43

9515.

1

KC8

2725

3.1

AY3

6224

3.1

KC8

3101

6.1

Dascyllus_ar

uanus

EF09

5632.

1

JQ43

1677.

1

JF458

114.1

KC8

2736

9.1

EF09

5674.

1

KC8

3117

4.1

Dascyllus_tri

maculatus

EU63

7953.

1

JQ43

1680.

1

JF458

123.1

EU6

3804

3.1

AY2

0865

5.1

EU2

5670

4.1

Diplodus_an

nularis

Y186

62.1

KC50

0532.

1

EU03

6432.

1

Dissostichus_

eleginoides

DQ4

9878

0.1

NC_0

18135

.1

NC_0

18135

.1

JN18

7039.

1

DQ4

9876

5.1

NC_0

18135

.1

NC_0

18135

.1

NC_0

18135

.1

NC_0

18135

.1

JX18

9902.

1

JN18

6957.

1

JN18

6821.

1

JX18

8975.

1

Drepane_afri

cana

AY1

4132

1.1

AY3

6224

4.1

EF09

5656.

1

EF61

7140.

1

Echeneis_na

ucrates

AY1

4131

5.1

KF02

1242.

1

KF02

1242.

1

AY3

6224

5.1

KF02

1242.

1

KF02

1242.

1

KF02

1242.

1

KF02

1242.

1

EU16

7829.

1

JX19

1011.

1

JX19

1334.

1

KF13

9499.

1

Elacatinus_m

acrodon

HQ5

3688

9.1

AF39

1348.

1

EU38

0935.

1

AF39

1420.

1

AF39

1492.

1

AY8

4655

2.1

HQ5

3679

3.1

Electrona_an

tarctica

AY1

4125

8.1

AP01

2248.

1

AP01

2248.

1

KF13

9987.

1

AY3

6220

1.1

AP01

2248.

1

AP01

2248.

1

AP01

2248.

1

AP01

2248.

1

KF14

1228.

1

KF14

1462.

1

KF14

0531.

1

Eleginops_m

aclovinus

AY1

41303.1

DQ52

6429.1

DQ52

6429.1

EU6

38047.1

DQ52

6429.1

DQ52

6429.1

DQ52

6429.1

DQ52

6429.1

KF14

1229.1

JN18

6941.1

JN18

6805.1

KF13

9574.1

Embiotoca_j

acksoni

EF09

5628.

JQ69

4028.

AF15

9331.

JX18

9716.

EF09

5670.

AY2

7988

JX18

9465.

JX18

9100.

JX18

8943.

7

1 1 1 1 1 2.1 1 1 1

Enchelyopus

_cimbrius

EU637958.

1

KF929836.

1

EU224006.

1

KC82720

6.1

FJ215237.

1

KC83079

3.1

Enoplosus_ar

matus

JF913270.

1

AP006008.

1

AP006008.

1

JQ352827.

1

AP006008.

1

AP006008.

1

AP006008.

1

AP006008.

1

EU167830.

1

JQ353139.

1

KF139501.

1

Epigonus_tel

escopus

EU637959.

1

HM007703

.1

DQ197949.

1

EU63804

8.1

EU167904.

1

KC83135

1.1

Epinephelus_

aeneus

AY14129

1.1

KC500672.

1

DQ197950.

1

HM05004

6.1

EU63804

9.1

HM050103

.1

HM04998

6.1

Eustomias_fil

ifer

KC311789.

1

GQ860382.

1

GQ86048

9.1

GQ86033

1.1

GQ86044

5.1

Euthynnus_a

lletteratus

DQ8

7480

7.1

AB09

9716.

1

AB09

9716.

1

HM0

5004

8.1

AB09

9716.

1

AB09

9716.

1

AB09

9716.

1

AB09

9716.

1

DQ8

7477

5.1

HM0

4999

0.1

Fistularia_pe

timba

AY1

4132

4.1

FJ541

073.1

JX19

0482.

1

JX19

0871.

1

JX19

1004.

1

KF13

9436.

1

Gadus_morh

ua

AF38

5832.

1

HG51

4359.

1

HG51

4359.

1

EU00

1906.

1

EU6

3805

0.1

HG51

4359.

1

HG51

4359.

1

HG51

4359.

1

HG51

4359.

1

KC4

4220

0.1

EU00

2070.

1

EU00

1871.

1

EU00

2017.

1

Gaidropsaru

s_biscayensis

KC98

1044.

1

EF42

7562.

1

Gempylus_se

rpens

DQ8

7481

2.1

AP01

2502.

1

AP01

2502.

1

KC8

2700

6.1

AP01

2502.

1

AP01

2502.

1

AP01

2502.

1

AP01

2502.

1

DQ8

7478

0.1

KC8

3110

1.1

Gnathanodo

n_speciosus

EU63

7963.

1

EF60

9362.

1

JQ93

9556.

1

EU6

3805

3.1

JQ93

8279.

1

JQ94

0161.

1

KC8

3123

8.1

Gobiodon_hi

strio

HQ5

3689

5.1

FJ583

450.1

KF41

5569.

1

AF39

1432.

1

AF32

2984.

1

KF41

5771.

1

HQ5

3679

9.1

KF41

5976.

1

Gobionotothe

n_acuta

DQ4

9878

6.1

JN64

0645.

1

DQ49

8809.

1

JN18

7041.

1

DQ4

9877

1.1

JN18

6897.

1

JN18

6959.

1

JN18

6823.

1

Gonorynchus

_greyi

EU40

9632

.1

NC_0

04702

.1

NC_0

04702

.1

JX19

0409.

1

NC_0

04702

.1

NC_0

04702

.1

NC_0

04702

.1

NC_0

04702

.1

EU40

9606.

1

JX19

0938.

1

JX19

1265.

1

KF13

9348.

1

Gonostoma_e

longatum

JN41

2561.

1

FJ918

931.1

FJ91

8869.

1

FJ89

6444.

1

KF14

1476.

1

FJ90

6665.

1

GQ8

6041

4.1

Gunnellichth

ys_monostig

ma

HQ5

3689

8.1

JQ43

1780.

1

AB02

1256.

1

AF39

1445.

1

AF39

1517.

1

KF41

5793.

1

KF41

5997.

1

Haemulon_a

urolineatum

EF09

5619.

1

JQ84

0858.

1

HQ67

6702.

1

KF14

0008.

1

EF09

5661.

1

EU6

9755

3.1

HQ6

6718

8.1

KF14

0545.

1

KF13

9505.

1

Himantoloph

us_groenland

icus

EU63

7965.

1

AB28

2840.

1

AB28

2840.

1

EU6

3805

5.1

AB28

2840.

1

AB28

2840.

1

AB28

2840.

1

AB28

2840.

1

Hippocampu

s_ramulosus

AY3

6833

0.1

AY3

6221

6.1

Hippoglossus

_hippoglossu

s

AF15

6265.

1

NC_0

09709

.1

NC_0

09709

.1

NC_0

09709

.1

NC_0

09709

.1

NC_0

09709

.1

NC_0

09709

.1

FJ76

9824.

1

JQ94

0204.

1

Holacanthus

_ciliaris

AY1

4132

2.1

JQ84

1232.

1

KC84

5364.

1

KC8

2714

5.1

AY3

6221

4.1

KC8

3123

3.1

Holanthias_c

hrysostictus

AY1

41290.1

GU3

68854.1

AY3

62209.1

EF09

5645.1

GU3

68705.1

Hoplostethus

_mediterrane

AY1

4126

EU86

9821.

GU54

9922.

AY3

6226

EF09

5635.

8

us 4.1 1 1 7.1 1

Howella_bro

diei

EU637966.

1

EU148199.

1

GU36886

8.1

EU63805

6.1

GU36883

1.1

GU36871

9.1

Istigobius_ri

gilius

HQ53694

6.1

HQ536672.

1

HQ536742.

1

FJ796137.1

KF235456.

1

HQ53685

4.1

Istiophorus_

platypterus

DQ87480

8.1

AB470306.

1

AB470306.

1

JQ939543.

1

JQ939436.

1

AB470306.

1

AB470306.

1

AB470306.

1

AB470306.

1

JQ938273.

1

DQ87477

6.1

KC83082

7.1

Kali_macrur

a

AY14130

8.1

EU148218.

1

AY36222

4.1

Katsuwonus_

pelamis

DQ87480

6.1

AB101290.

1

AB101290.

1

AB101290.

1

AB101290.

1

AB101290.

1

AB101290.

1

DQ87477

4.1

KC83109

7.1

Lagocephalu

s_laevigatus

AY1

4128

5.1

NC_0

15345

.1

NC_0

15345

.1

KF02

7847.

1

AY3

6222

1.1

NC_0

15345

.1

NC_0

15345

.1

NC_0

15345

.1

NC_0

15345

.1

JQ68

2323.

1

KF02

8077.

1

KF02

8274.

1

KF02

7643.

1

Lampanyctus

_alatus

JN41

2575.

1

JX14

9658.

1

Lampris_im

maculatus

AY1

4125

9.1

DQ10

8066.

1

Lamprogram

mus_shcherb

achevi

EU63

7969.

1

EU6

3805

8.1

Leiognathus_

fasciatus

EU63

7972.

1

HQ14

9860.

1

EU6

3806

0.1

Liparis_fabri

cii

AY3

6831

7.1

KC01

5558.

1

EF50

8319.

1

AY3

6223

5.1

Liza_saliens Y186

70.1

JQ06

0483.

1

JQ06

0226.

1

Lophius_bud

egassa

EF09

5608.

1

JQ77

4652.

1

EF43

9543.

1

EF09

5637.

1

Lutjanus_seb

ae

EU63

7974.

1

NC_0

12737

.1

NC_0

12737

.1

EU6

3806

4.1

NC_0

12737

.1

NC_0

12737

.1

NC_0

12737

.1

NC_0

12737

.1

EU62

7668.

1

EU6

2765

6.1

Luvarus_imp

erialis

EU63

7975.

1

NC_0

09851

.1

NC_0

09851

.1

EF53

6299.

1

EU6

3806

5.1

NC_0

09851

.1

NC_0

09851

.1

NC_0

09851

.1

NC_0

09851

.1

EF53

0099.

1

KF14

1513.

1

EF53

3922.

1

EF53

9246.

1

Lycodapus_a

ntarcticus

EU63

7976.

1

JN64

0678.

1

EU6

3806

6.1

Macroramph

osus_scolopa

x

AY1

4128

0.1

NC_0

10265

.1

NC_0

10265

.1

JX19

0483.

1

AY3

6220

6.1

NC_0

10265

.1

NC_0

10265

.1

NC_0

10265

.1

NC_0

10265

.1

JX19

0872.

1

JX19

1005.

1

JX19

1329.

1

KF13

9438.

1

Makaira_nig

ricans

DQ8

7481

0.1

HM9

09858

.1

EF39

2596.

1

DQ85

4672.

1

EU16

7744.

1

DQ8

7477

8.1

Mene_macul

ata

AY1

4131

6.1

AB35

5909.

1

AB35

5909.

1

JQ93

9501.

1

AY3

6364

2.1

AB35

5909.

1

AB35

5909.

1

AB35

5909.

1

AB35

5909.

1

EF09

5659.

1

JX19

1020.

1

JX19

1341.

1

KF13

9516.

1

Menidia_me

nidia

EU63

7977.

1

NC_0

11174

.1

NC_0

11174

.1

EU6

3806

7.1

NC_0

11174

.1

NC_0

11174

.1

NC_0

11174

.1

NC_0

11174

.1

JQ28

2082.

1

KC8

3082

5.1

Menticirrhus

_americanus

DQ8

7482

1.1

KF93

0121.

1

JQ32

2699.

1

DQ8

7478

9.1

Merlangius_

merlangus

AY1

41260.1

NC_0

07395.1

NC_0

07395.1

KC8

27213.1

NC_0

07395.1

NC_0

07395.1

NC_0

07395.1

NC_0

07395.1

FJ21

5265.1

Microchirus_

variegatus

AY1

4128

JQ77

5080.

EF43

9551.

AY3

6227

9

4.1 1 1 5.1

Microdesmus

_longipinnis

HQ53688

5.1

AF391341.

1

EU380926.

1

AF391414.

1

AF391486.

1

KC83075

0.1

Mola_mola AF137215.

1

NC_005836

.1

NC_005836

.1

KF027832.

1

AY36225

1.1

NC_005836

.1

NC_005836

.1

NC_005836

.1

NC_005836

.1

EF095643.

1

KF028062.

1

JX191355.

1

KF027627.

1

Mullus_surm

uletus

EU637982.

1

KC500992.

1

DQ197965.

1

AY36223

1.1

EF095658.

1

Myripristis_

berndti

U57538.1

AP002940.

1

AP002940.

1

JX390835.

1

AP002940.

1

AP002940.

1

AP002940.

1

AP002940.

1

Myripristis_

murdjan

KC442231.

1

FJ459546.1

JX390842.

1

JX390911.

1

KF140576.

1

JX390762.

1

Myripristis_v

iolacea

U575

39.1

FJ583

677.1

EU00

1916.

1

JX39

0914.

1

EU00

2078.

1

EU00

1880.

1

EU00

2026.

1

Naso_liturat

us

EU63

7984.

1

JQ35

0127.

1

AB27

6964.

1

EF53

6292.

1

EU6

3807

4.1

EF53

0092.

1

KF14

1532.

1

EF53

3915.

1

EF53

9239.

1

Nemateleotri

s_magnifica

HQ5

3688

6.1

JQ35

0136.

1

EU38

0918.

1

AF39

1400.

1

AF39

1472.

1

KF14

1288.

1

HQ5

3679

2.1

KF14

0579.

1

Neomerinthe

_hemingwayi

DQ8

7481

9.1

KC8

2717

1.1

DQ8

7478

7.1

KC8

3112

1.1

Neoniphon_a

rgenteus

U575

40.1

JX39

0737.

1

U575

27.1

JX39

0846.

1

JX39

0916.

1

KC8

3122

9.1

JX39

0767.

1

Neoniphon_s

ammara

U575

36.1

KC18

7866.

1

KC18

8040.

1

JX39

0849.

1

JX39

0919.

1

KC8

3138

4.1

JX39

0770.

1

Niphon_spin

osus

EU63

7934.

1

EF14

3386.

1

DQ27

4053.

1

HQ7

3135

6.1

JQ08

8491.

1

HQ7

3131

3.1

KF14

1537.

1

HQ7

3118

4.1

KF13

9555.

1

Notothenia_c

oriiceps

AY1

4130

2.1

NC_0

15653

.1

NC_0

15653

.1

HM0

5006

3.1

AY3

6365

5.1

NC_0

15653

.1

NC_0

15653

.1

NC_0

15653

.1

NC_0

15653

.1

EU16

7892.

1

JN18

6965.

1

JN18

6829.

1

HM0

5000

3.1

Oplopomus_

oplopomus

HQ5

3693

6.1

HQ53

6732.

1

FJ796

151.1

KF23

5472.

1

HQ5

3684

2.1

KC8

3077

0.1

Opsanus_par

dus

DQ8

7482

2.1

AF16

5347.

1

JX19

0479.

1

JX19

0866.

1

DQ8

7479

0.1

JX19

0999.

1

KF13

9407.

1

Ostracion_cu

bicus

KF02

7990.

1

JQ35

0150.

1

EU27

4416.

1

KF02

7842.

1

KF0

2780

0.1

KF02

7953.

1

KF02

8073.

1

JX18

9160.

1

KF02

7638.

1

Pagothenia_b

orchgrevinki

DQ4

9879

1.1

GU99

7399.

1

EU25

0417.

1

HM1

6636

5.1

DQ4

9877

7.1

DQ18

4489.

1

HM1

6623

9.1

HM1

6619

0.1

Paradiplospi

nus_antarctic

us

DQ8

7481

4.1

EU26

3805.

1

EU26

3851.

1

DQ8

7478

2.1

Paralichthys

_olivaceus

HQ4

1377

2.1

NC_0

02386

.1

NC_0

02386

.1

NC_0

02386

.1

NC_0

02386

.1

NC_0

02386

.1

NC_0

02386

.1

KC4

4221

0.1

Patagonototh

en_tessellata

DQ4

9878

2.1

EU07

4538.

1

DQ49

8805.

1

HM1

6636

3.1

DQ4

9876

7.1

AY25

6570.

1

HM1

6628

8.1

HM1

6623

8.1

HM1

6618

8.1

Pholis_gunne

llus

AY1

4129

8.1

KC01

5798.

1

AY3

6228

5.1

Photostomias

_guernei

JN41

2566.1

GQ86

0402.1

GQ8

60509.1

GQ8

60353.1

GQ8

60465.1

Phycis_blenn

oides

AY3

6832

JQ77

4694.

DQ17

4073.

10

1.1 1 1

Polymixia_no

bilis

AY36832

0.1

DQ197980.

1

HM05007

3.1

AY36220

8.1

HM050133

.1

HM05001

5.1

Pomacanthus

_maculosus

EU637995.

1

HQ149906.

1

JN604381.

1

EU63807

9.1

EU167864.

1

Pomacentrus

_pavo

EF095631.

1

JF435124.1

JF458235.1

KC82736

6.1

EF095673.

1

JQ707284.

1

KC83116

0.1

Pontinus_lon

gispinis

EU637996.

1

KC82717

2.1

EU63808

1.1

KC83122

2.1

Priacanthus_

arenatus

EU637997.

1

KF930309.

1

KF140090.

1

EU63808

2.1

KF141327.

1

KF141575.

1

KF140607.

1

KF139547.

1

Priolepis_cin

cta

HQ5

3690

1.1

JQ35

0252.

1

KC8

2692

1.1

AF39

1457.

1

AF39

1529.

1

HQ5

3680

5.1

Psettodes_bel

cheri

JQ93

8077.

1

JQ93

9639.

1

AY3

6225

9.1

JQ94

0224.

1

Psettodes_er

umei

KC44

2235.1

NC_0

20032

.1

NC_0

20032

.1

JQ93

9638.

1

JQ93

9480.

1

NC_0

20032

.1

NC_0

20032

.1

NC_0

20032

.1

NC_0

20032

.1

JQ93

8325.

1

JQ94

0223.

1

JX19

1347.

1

Pseudopleuro

nectes_ameri

canus

AY6

31036.1

KF93

0333.1

JX18

9771.1

FJ870

451.1

JX18

9931.1

FJ87

0490.1

JX18

9523.1

JX18

9157.1

JX18

9002.1

Ptereleotris_

microlepis

HQ5

36975.1

AF39

1380.1

EU38

0951.1

HQ90

9501.1

AF39

1524.1

KF23

5479.1

HQ9

09608.1

KC8

30758.1

Pterycombus

_brama

EU63

8001.1

GU22

5018.1

EU6

38086.1

KC8

31065.1

Regalecus_gl

esne

AY3

68328.1

FJ918

947.1

JF931

967.1

KF14

0111.1

AY3

62266.1

JF93

1987.1

KF14

1593.1

EU00

1874.1

Risor_ruber HQ5

36890.1

AF39

1352.1

KF41

5655.1

AF39

1424.1

AF39

1495.1

KF41

5859.1

HQ5

36794.1

KF41

6067.1

Ruvettus_pre

tiosus

DQ8

74813.1

NC_0

22493.1

NC_0

22493.1

JX18

9762.1

NC_0

22493.1

NC_0

22493.1

NC_0

22493.1

NC_0

22493.1

JX18

9922.1

DQ8

74781.1

JX18

9516.1

JX18

9149.1

JX18

8993.1

Rypticus_sap

onaceus

AY3

68329.1

JN82

8108.1

AY32

1800.1

JX18

9703.1

AY3

62257.1

JX18

9861.1

JX18

9617.1

JX18

9087.1

JX18

8928.1

Sarda_sarda DQ8

74800.1

KC50

1212.1

EU03

6483.1

JX18

9763.1

EU26

3833.1

JX18

9923.1

DQ8

74768.1

JX18

9517.1

JX18

9150.1

JX18

8994.1

Sargocentron

_diadema

U575

37.1

JQ35

0312.1

U575

30.1

KC8

27338.1

JX39

0778.1

Sargocentron

_tiere

U575

45.1

JQ35

0323.1

U575

34.1

JX39

0867.1

JX39

0934.1

JX39

0788.1

Sarpa_salpa Y186

64.1

KC50

1272.1

DQ19

7992.1

JQ93

9515.1

HQ6

76686.1

JQ94

0134.1

KC8

31092.1

Scarus_psitta

cus

EF09

5633.1

JQ43

2113.1

EU60

1387.1

EF09

5675.1

EU6

01337.1

Scomber_jap

onicus

AY1

41311.1

AB10

2724.1

AB10

2724.1

AY3

62237.1

AB10

2724.1

AB10

2724.1

AB10

2724.1

AB10

2724.1

KC8

31102.1

Scomberomo

rus_maculat

us

DQ87479

8.1

DQ835941.

1

EU349362.

1

KF140119.

1

AF076128.

1

KF141356.

1

DQ87476

6.1

KF141602.

1

KF140636.

1

KF139608.

1

11

Scomber_sco

mbrus

DQ8

7479

7.1

NC_0

06398

.1

NC_0

06398

.1

KF14

0118.

1

NC_0

06398

.1

NC_0

06398

.1

NC_0

06398

.1

NC_0

06398

.1

EU47

7493.

1

DQ8

7476

5.1

JQ94

0142.

1

KF14

0635.

1

KF13

9607.

1

Scophthalmu

s_rhombus

EU63

8005.

1

EU51

3711.

1

EU03

6491.

1

KF31

2005.

1

Scorpaena_el

ongata

EF45

6081.

1

EF45

6020.

1

Sebastes_alut

us

EF21

2407.

1

JQ35

4342.

1

AB09

6111.

1

HQ12

4007.

1

DQ6

7871

8.1

Sebastolobus

_altivelis

DQ4

9012

4.1

JQ35

4499.

1

Selar_crume

nophthalmus

DQ8

7482

0.1

HQ56

1001.

1

AF51

5743.

1

JQ93

9561.

1

JQ93

9450.

1

DQ8

7478

8.1

JQ94

0164.

1

KC8

3135

5.1

Selene_dorsa

lis

EU63

8006.

1

AF36

3744.

1

EU6

3808

9.1

Serranus_acc

raensis

AY1

4128

9.1

AY3

6220

2.1

Siganus_vulp

inus

EU63

8007.1

FJ584

118.1

AY19

0548.1

EF53

6306.1

EU6

38090.1

EF53

0105.1

JX19

1023.1

EF53

3929.1

KF13

9602.1

Sillago_siha

ma

EU63

8008.1

NC_0

16672.1

NC_0

16672.1

JQ93

9514.1

NC_0

16672.1

NC_0

16672.1

NC_0

16672.1

NC_0

16672.1

EU16

7874.1

KF14

1610.1

Solea_solea EU63

8009.1

KJ20

5196.1

DQ19

8003.1

EF09

5644.1

Sphaeramia_

nematoptera

EU63

8010.1

FJ584

123.1

EU6

38091.1

Sphyraena_b

arracuda

DQ8

74816.1

NC_0

22484.1

NC_0

22484.1

JQ93

9563.1

JQ93

9453.1

NC_0

22484.1

NC_0

22484.1

NC_0

22484.1

NC_0

22484.1

JQ93

8282.1

DQ8

74784.1

JQ94

0167.1

JX19

1346.1

KF13

9610.1

Sphyraena_s

phyraena

AY1

41312.1

KJ70

9637.1

EF43

9597.1

AY3

62254.1

Spinachia_sp

inachia

AY1

41281.1

NC_0

11582.1

NC_0

11582.1

AB4

45157.1

AB4

45139.1

NC_0

11582.1

NC_0

11582.1

NC_0

11582.1

NC_0

11582.1

AB4

45184.1

AB4

45193.1

AB4

45220.1

Stomias_boa JN41

2565.1

KF93

0474.1

EU00

1914.1

EU00

2076.1

EU00

1879.1

EU00

2024.1

Stonogobiops

_xanthorhini

ca

HQ5

36930.1

HQ53

6655.1

HQ53

6727.1

FJ796

152.1

HQ5

36836.1

Syacium_mic

rurum

AY3

68334.1

KF93

0479.1

JQ93

9599.1

AY3

62262.1

JQ94

0199.1

Synagrops_b

ellus

JF91

3271.1

EU18

2640.1

AF12

3592.1

EU18

2622.1

EU18

2628.1

EU18

2646.1

Synanceia_ve

rrucosa

EU63

8011.1

JQ43

2179.1

KC8

26980.1

EU6

38093.1

KC8

31340.1

Syngnathus_t

yphle

AY3

68326.1

AF35

6042.1

AY3

62211.1

Synodus_foet

ens

DQ8

74823.1

JX51

9411.1

EU00

1918.1

JN23

0900.1

DQ8

74791.1

EU00

2080.1

JX19

1302.1

EU00

2028.1

Thunnus_atl

anticus

DQ8

74802.1

JQ36

5595.1

AB09

8104.1

DQ8

74770.1

12

Trachinotus_

ovatus

AY1

4131

4.1

NC_0

22707

.1

NC_0

22707

.1

JQ93

9547.

1

AY3

6226

3.1

NC_0

22707

.1

NC_0

22707

.1

NC_0

22707

.1

NC_0

22707

.1

KF14

1380.

1

JQ94

0152.

1

KF14

0654.

1

KF13

9479.

1

Trachurus_tr

achurus

EU63

8013.

1

NC_0

06818

.1

NC_0

06818

.1

NC_0

06818

.1

NC_0

06818

.1

NC_0

06818

.1

NC_0

06818

.1

KF31

1975.

1

Triacanthode

s_anomalus

KC44

2243.

1

AP00

9172.

1

AP00

9172.

1

EF53

6312.

1

EU6

3809

5.1

AP00

9172.

1

AP00

9172.

1

AP00

9172.

1

AP00

9172.

1

AY3

0878

8.1

JX18

9529.

1

EF53

3935.

1

EF53

9259.

1

Triacanthode

s_ethiops

KF02

8008.

1

JQ86

1002.

1

KF02

7576.

1

KF02

7860.

1

KF0

2781

8.1

KF02

7967.

1

KF02

8090.

1

KF02

8286.

1

KF02

7654.

1

Trichiurus_le

pturus

DQ8

7479

6.1

NC_0

18791

.1

NC_0

18791

.1

JX18

9764.

1

NC_0

18791

.1

NC_0

18791

.1

NC_0

18791

.1

NC_0

18791

.1

EU16

7903.

1

DQ8

7476

4.1

JX18

8995.

1

Uranoscopus

_albesca

AY1

4130

5.1

AY3

6223

9.1

EU16

7748.

1

Valenciennea

_puellaris

HQ5

3691

0.1

FJ584

213.1

HQ53

6710.

1

FJ796

157.1

HQ5

3681

4.1

Valenciennea

_strigata

EU63

8017.

1

JQ43

2215.

1

EU38

0910.

1

KC8

2690

5.1

AF39

1456.

1

AF39

1528.

1

AB42

9399.

1

KF41

5885.

1

HQ5

3680

4.1

KF41

6095.

1

Vanderhorsti

a_ornatissim

a

HQ5

36967.1

HQ53

6690.1

KF41

5680.1

KC8

26912.1

HQ53

6764.1

HQ53

6790.1

KF41

5886.1

HQ5

36876.1

KF41

6096.1

Xeneretmus_

latifrons

EU63

8018.1

FJ165

462.1

KC8

27136.1

EU6

38097.1

KC8

31324.1

Xiphias_glad

ius

EU63

8019.1

AP00

6036.1

AP00

6036.1

JQ93

9568.1

EU6

38098.1

AP00

6036.1

AP00

6036.1

AP00

6036.1

AP00

6036.1

KF31

1985.1

DQ8

74779.1

JQ94

0177.1

JX19

1365.1

KF13

9613.1

Xyrichtys_no

vacula

EU63

8020.1

JQ83

9672.1

EF43

9247.1

KC8

27348.1

Zenopsis_con

chifer

AY3

68314.1

KC01

6043.1

KF14

0150.1

AY3

62286.1

AY3

08778.1

KF14

1634.1

KF13

9385.1

Zeus_faber EU63

8023.1

NC_0

03190.1

NC_0

03190.1

JQ93

9540.1

AF1

37241.1

NC_0

03190.1

NC_0

03190.1

NC_0

03190.1

NC_0

03190.1

JN23

0905.1

JX19

0988.1

EU00

1894.1

KF13

9386.1

13

14

15

16

17

18

Figure S1. Chronogram used in this study, adapted from Betancur et al. 2013, showing the

relationship for 194 euteleosts and the two outgroup species Alepocephalus bairdii and

Gonorynchus greyi. Branch lengths are in units of millions of years.

Figure S2. Snake plot of rhodopsin labeling sites which correlated with habitat depth in black.

E1-E3: Extracellular loops 1 to 3, C1-C3: Cytoplasmic loops 1 to 3, NT: N-terminus, CT: C-

terminus, H1-H8: helices 1 to 8, disulfide bond between C110 and C187

19

Figure S3. Box-plots showing the difference in median habitat depths of species with different

residues at some of the RH1 sites correlating with habitat depth.

0

500

1000

1500

2000

Dep

th (

m)

A114 G114

0

500

1000

1500

2000

Dep

th (

m)

A144 S144

C185 V185 A166 S166 T166

20

(I a)

(I b)

KW

PGLS

21

(I c)

(I d)

DISC

AR

22

KW

PGLS (II b)

(II a)

23

DISC

AR (II d)

(II c)

24

(III a)

(III b) PGLS

KW

25

(III d)

(III c) DISC

AR

26

KW

PGLS

(IV a)

(IV b)

27

AR

DISC (IV c)

(IV d)

28

(V b)

(V a)

PGLS

KW

29

(V d)

(V c) DISC

AR

30

PGLS

KW

(VI b)

(VI a)

31

AR

DISC (VI c)

(VI d)

32

(VII b)

(VII a) KW

PGLS

33

(VII d)

(VII c)

AR

DISC

34

KW

PGLS (VIII b)

(VIII a)

35

DISC

AR

(VIII c)

(VIII d)

36

PGLS

KW

(IX b)

(IX a)

37

AR

DISC (IX c)

(IX d)

38

PGLS

KW

(X b)

(X a)

39

AR

DISC (X c)

(X d)

40

PGLS

KW

(XI b)

(XI a)

41

AR

DISC (XI c)

(XI d)

42

KW

PGLS

(XII a)

(XII b)

43

AR

DISC (XII c)

(XII d)

44

PGLS

KW (XIII a)

(XIII b)

45

Figure S4. Distribution of p-values for the sites in rhodopsin compared to the control genes used

in this study. RH1 data set was pruned to only include the sequences for which a corresponding

control gene sequence was available in order to eliminate the effect of sample size difference in

comparisons. Only sites for which a second residue identity could be found in more than five

sequences are shown here. KW: Kruskal-Wallis, PGLS: Phylogenetic Generalized Least

Squares, DISC: Discrete, AR: Ancestral Reconstructions. RH1 was compared to the following

control genes: (I) COI, (II) CYTB, (III) RAG1, (IV) MYH6, (V) ZIC1, (VI) MLL, (VII) ND2,

(VIII) ND1, (IX) SH3PX3, (X) ENC1, (XI) ND4, (XII) RAG2, (XIII) ND5

AR

DISC (XIII c)

(XIII d)

46

Chapter 3: Investigating Correlations between habitat water

temperature of teleost fishes and amino acid substitutions in

their rhodopsins

47

48

49

50

Figure S1. Literature generated species tree showing the relationships between 105

ostariophysan fishes and the two outgroups Lepisosteus oculatus and Osteoglossum bicirrhosum.

Branch lengths are in units of nucleotide substitutions per codon sites obtained from M8 model

on RH1 sequences.