水的深度及水中懸浮微粒的多寡等因素使環境光譜分佈迥然不同，光從清澈的海水表層向下穿透，光環境隨著深度而由全光譜分佈變成只剩下藍光，而在混濁的水域中，短波長的藍光容易被懸浮微粒吸收，使紅光變成光環境中優勢的光波長。鰕虎科魚類棲息的環境廣泛，遍布不同光環境中的，適合作為研究光環境變化與視覺基因演化之關係的物種。本研究選擇三種不同棲地的鰕虎科魚類作為研究的對象，包含(1) 海水域的縱帶鸚鰕虎 (Exyrias puntang)；(2) 河口泥灘地的彈塗魚 (Periophthalmus modestus)；(3) 淡水流域的道氏短鰕虎 (Brachygobius doriae)，比較其視覺基因種類、數量及胺基酸序列上的差異。結果顯示，三種鰕虎的視覺基因除了數量上的差異之外，視覺基因的光譜調節位點也有差別：彈塗魚及道氏短鰕虎都擁有SWS2A/B兩個短波長基因，而且這兩個基因之間有L116T的胺基酸替換。三個物種都擁有LWSA/B基因，道氏短鰕虎更擁有LWSA1/A2及LWSB共三個長波長基因；在本研究中LWSA/B兩個基因之間有Y261F的差異，彈塗魚的LWSB基因及道氏短鰕虎的LWSA2及LWSB基因則有S164A的胺基酸替換，且各個LWS基因的S164A胺基酸替換是演化過程中個別發生的。視覺基因的變異在演化過程中日積月累，其中LWSA/B基因複製事件發生在真骨魚類的共同祖先出現之前，但只有鰕虎科魚類保留LWSB基因，推測辨識紅光對鰕虎科魚類可能相當重要。由本研究的結果推論，基因複製及光譜調節位點的胺基酸替換在演化過程中發生，使三種鰕虎對於各色光波段的感光能力並不一樣；棲地環境相對複雜的彈塗魚及道氏短鰕虎擁有更好的藍光感知能力，而道氏短鰕虎棲息在紅色光譜分佈易改變的淡水環境，則擁有更好的紅光感知能力，顯示不同棲地物種的視覺基因受到光環境影響而存在分歧。

Water depth and the amount of suspended particles shape the light spectrum of an aquatic environment. In clean open water, the full spectrum is attenuated to blue right as the depth increases. However, shortwave blue light is absorbed by suspended particles in turbid water, leaving a red light dominant photic environment. Gobiidae are distributed widely in various habitats with varying photic environments, making them good candidates to study the relationship of habitat divergence and fish opsin evolution. Three goby species, freshwater bumblebee goby (Brachygobius doriae), brackish mudskipper (Periophthalmus modestus) and ocean silver-spotted goby (Exyrias puntang), inhabiting different photic environments were used to study the relationship between opsin gene evolution and habitat divergences. Two types of SWS2 genes, SWS2A and 2B, were identified in these gobies, and an amino acid substitution L116T, were found between them. Three types of LWS genes were observed in this study, LWSA1/A2 and LWSB. Two substitutions, A164S and F261Y, were found among LWS genes. These substitutions make mudskippers and bumblebee gobies better ability to detect blue or blue/red light, respectively. We also found gene duplication events occurred twice in LWS, and LWSA/B gene duplicated before common ancestor of teleost appeared. Futhermore, only gobies in this study possess LWSB gene while other teleost loss it and this is inferring that detecting red light might quite nececery for gobiidae. All these differences appeared among evolution and generating a diverse set of opsin gene of gobies living in different habitats. This result suggest opsin genes of species in this study were distinct because of affect from photic environments.

論文審定書 i
論文公開授權書 ii
謝辭 iii
中文摘要 iv
英文摘要 v
目錄 vi
表目錄 vii
圖目錄 viii
附錄 ix
壹、前言 1
貳、材料與方法 8
參、結果 15
肆、討論 20
伍、結論 28
參考文獻 29
附表 36
附圖 43
附錄 48

Archer, S., A. Hope, and J. C. Partridge. 1995. The molecular basis for the green-blue sensitivity shift in the rod visual pigments of the European eel. Proceedings of the Royal Society B: Biological Sciences 262(1365):289-295.
Archer, S. N., and J. N. Lythgoe. 1990. The visual pigment basis for cone polymorphism in the guppy, Poecilia reticulata. Vision Research 30(2):225-233.
Archer, S. N., J. N. Lythgoe, and L. Hall. 1992. Rod opsin cDNA sequence from the sand goby (Pomatoschistus minutus) compared with those of other vertebrates. Proceedings of the Royal Society B: Biological Sciences 248(1321):19-25.
Asenjo, A. B., J. Rim, and D. D. Oprian. 1994. Molecular determinants of human red/green color discrimination. Neuron 12(5):1131-1138.
Bailes, H. J., W. L. Davies, A. E. Trezise, and S. P. Collin. 2007. Visual pigments in a living fossil, the Australian lungfish Neoceratodus forsteri. BMC Evolutionary Biology 7(1):200.
Bowmaker, J. K. 1995. The visual pigments of fish. Progress in Retinal and Eye Research 15(1):1-31.
Bowmaker, J. K. 2008. Evolution of vertebrate visual pigments. Vision Research 48(20):2022-2041.
Carleton, K. L., F. I. Harosi, and T. D. Kocher. 2000. Visual pigments of African cichlid fishes: evidence for ultraviolet vision from microspectrophotometry and DNA sequences. Vision Research 40(8):879-890.
Carleton, K. L., and T. D. Kocher. 2001. Cone opsin genes of african cichlid fishes: tuning spectral sensitivity by differential gene expression. Molecular Biology and Evolution 18(8):1540-1550.
Carleton, K. L., J. W. Parry, J. K. Bowmaker, D. M. Hunt, and O. Seehausen. 2005. Colour vision and speciation in Lake Victoria cichlids of the genus Pundamilia. Molecular Ecology 14(14):4341-4353.
Chen, W. J., C. Bonillo, and G. Lecointre. 2003. Repeatability of clades as a criterion of reliability: a case study for molecular phylogeny of Acanthomorpha (Teleostei) with larger number of taxa. Molecular Phylogenetics and Evolution 26(2):262-288.
Chinen, A., T. Hamaoka, Y. Yamada, and S. Kawamura. 2003. Gene duplication and spectral diversification of cone visual pigments of zebrafish. Genetics 163(2):663-675.
Chinen, A., Y. Matsumoto, and S. Kawamura. 2005. Spectral differentiation of blue opsins between phylogenetically close but ecologically distant goldfish and zebrafish. Journal of Biological Chemistry 280(10):9460-9466.
Cowing, J. A., S. Poopalasundaram, S. E. Wilkie, J. K. Bowmaker, and D. M. Hunt. 2002. Spectral tuning and evolution of short wave-sensitive cone pigments in cottoid fish from Lake Baikal. Biochemistry 41(19):6019-6025.
Davies, W. L., L. S. Carvalho, B. H. Tay, S. Brenner, D. M. Hunt, and B. Venkatesh. 2009. Into the blue: gene duplication and loss underlie color vision adaptations in a deep-sea chimaera, the elephant shark Callorhinchus milii. Genome Research 19(3):415-426.
Felsenstein, J. 1985. Confidence-limits on phylogenies - an approach using the bootstrap. Evolution 39(4):783-791.
Frohman, M. A., M. K. Dush, and G. R. Martin. 1988. Rapid production of full-length cdnas from rare transcripts - amplification using a single gene-specific oligonucleotide primer. Proceedings of the National Academy of Sciences of the United States of America 85(23):8998-9002.
Goldman, N., and Z. Yang. 1994. A codon-based model of nucleotide substitution for protein-coding DNA sequences. Molecular Biology and Evolution 11(5):725-736.
Graur, D., and W. H. Li. 2000. Fundamentals of Molecular Evolution. 2nd edition. Sinauer Associates, Sunderland, Mass.
Hamaoka, T., M. Takechi, A. Chinen, Y. Nishiwaki, and S. Kawamura. 2002. Visualization of rod photoreceptor development using GFP-transgenic zebrafish. Genesis: The Journal of Genetics and Development 34(3):215-220.
Hasegawa, M., H. Kishino, and T. Yano. 1985. Dating of the human-ape splitting by a molecular clock of mitochondrial DNA. Journal of Molecular Evolution 22(2):160-174.
Hisatomi, O., S. Kayada, Y. Aoki, T. Iwasa, and F. Tokunaga. 1994. Phylogenetic-relationships among vertebrate visual pigments. Vision Research 34(23):3097-3102.
Hofmann, C. M., and K. L. Carleton. 2009. Gene duplication and differential gene expression play an important role in the diversification of visual pigments in fish. Integr Comp Biol 49(6):630-643.
Hofmann, C. M., N. J. Marshall, K. Abdilleh, Z. Patel, U. E. Siebeck, and K. L. Carleton. 2012. Opsin evolution in damselfish: convergence, reversal, and parallel evolution across tuning sites. Journal of Molecular Evolution 75(3-4):79-91.
Horn, M. H., K. L. Martin, and M. A. Chotkowski. 1998. Intertidal fishes: life in two worlds. Academic Press, San Diego, California.
Hunt, D. M., K. S. Dulai, J. C. Partridge, P. Cottrill, and J. K. Bowmaker. 2001. The molecular basis for spectral tuning of rod visual pigments in deep-sea fish. Journal of Experimental Biology 204(Pt 19):3333-3344.
Hunt, D. M., J. Fitzgibbon, S. J. Slobodyanyuk, and J. K. Bowmaker. 1996. Spectral tuning and molecular evolution of rod visual pigments in the species flock of cottoid fish in Lake Baikal. Vision Research 36(9):1217-1224.
Jackson, D., F. Lewis, G. Taylor, A. Boylston, and P. Quirke. 1990. Tissue extraction of DNA and RNA and analysis by the polymerase chain reaction. Journal of Clinical Pathology 43(6):499-504.
Jerlov, N. G. 1968. Scattering. Pages 15-46 in Optical Oceanography. Elsevier, Netherlands.
Johnson, R. L., K. B. Grant, T. C. Zankel, M. F. Boehm, S. L. Merbs, J. Nathans, and K. Nakanishi. 1993. Cloning and expression of goldfish opsin sequences. Biochemistry 32(1):208-214.
Jokela-Maatta, M., A. Vartio, L. Paulin, and K. Donner. 2009. Individual variation in rod absorbance spectra correlated with opsin gene polymorphism in sand goby (Pomatoschistus minutus). Journal of Experimental Biology 212(Pt 21):3415-3421.
Jokela-Maatta, M., A. Vartio, L. Paulin, N. Fyhrquist-Vanni, and K. Donner. 2003. Polymorphism of the rod visual pigment between allopatric populations of the sand goby (Pomatoschistus minutus): a microspectrophotometric study. Journal of Experimental Biology 206(15):2611-2617.
Kimura, M. 1980. A simple method for estimating evolutionary rates of base substitutions through comparative studies of nucleotide sequences. Journal of Molecular Evolution 16(2):111-120.
Kirk, J. T. O. 1994. Scattering of light within the aquatic medium. Pages 98-132 in Light and photosynthesis in aquatic ecosystems, 2nd edition. Cambridge University Press, New York.
Larmuseau, M. H., T. Huyse, K. Vancampenhout, J. K. Van Houdt, and F. A. Volckaert. 2010. High molecular diversity in the rhodopsin gene in closely related goby fishes: A role for visual pigments in adaptive speciation? Molecular Phylogenetics and Evolution 55(2):689-698.
Larmuseau, M. H., M. P. Vanhove, T. Huyse, F. A. Volckaert, and R. Decorte. 2011. Signature of selection on the rhodopsin gene in the marine radiation of American seven-spined gobies (Gobiidae, Gobiosomatini). Journal of Evolutionary Biology 24(7):1618-1625.
Le, S. Q., and O. Gascuel. 2008. An improved general amino acid replacement matrix. Molecular Biology and Evolution 25(7):1307-1320.
Locket, N. A. 1971. Retinal structure in Platytroctes apus, a deep-sea fish with a pure rod fovea. Journal of the Marine Biological Association of the United Kingdom 51(01):79-91.
Locket, N. A. 1977. Adaptation to the deep-sea environment. Pages 67-192 in H. Autrum, R. Jung, W. R. Loewenstein, D. M. MacKay, and H. L. Teuber, editors. Handbook of Sensory Physiology, volume VII. Springer, Berlin.
Matsumoto, Y., S. Fukamachi, H. Mitani, and S. Kawamura. 2006. Functional characterization of visual opsin repertoire in Medaka (Oryzias latipes). Gene 371(2):268-278.
Mol, J. H. 2012a. The freshwater fishes of Suriname. Pages 56-62, volume 2. Brill, Leiden, Netherlands.
Mol, J. H. 2012b. The freshwater fishes of Suriname. Pages 35-50, volume 2. Brill, Leiden, Netherlands.
Munz, F. W. 1958a. Photosensitive pigments from the retinae of certain deep-sea fishes. Journal of General Physiology 140(2):220-235.
Munz, F. W. 1958b. The photosensitive retinal pigments of fishes from relatively turbid coastal waters. Journal of General Physiology 42(2):445-459.
Nagai, H., Y. Terai, T. Sugawara, H. Imai, H. Nishihara, M. Hori, and N. Okada. 2011. Reverse evolution in RH1 for adaptation of cichlids to water depth in Lake Tanganyika. Molecular Biology and Evolution 28(6):1769-1776.
Nathans, J. 1990. Determinants of visual pigment absorbance: role of charged amino acids in the putative transmembrane segments. Biochemistry 29(4):937-942.
Nawrocki, L., R. BreMiller, G. Streisinger, and M. Kaplan. 1985. Larval and adult visual pigments of the zebrafish, Brachydanio rerio. Vision Research 25(11):1569-1576.
Nei, M., and S. Kumar. 2000. Molecular evolution and phylogenetics. Oxford University Press, New York.
Nielsen, R., and Z. Yang. 1998. Likelihood models for detecting positively selected amino acid sites and applications to the HIV-1 envelope gene. Genetics 148(3):929-936.
Oprian, D. D., R. S. Molday, R. J. Kaufman, and H. G. Khorana. 1987. Expression of a synthetic bovine rhodopsin gene in monkey kidney cells. Proceedings of the National Academy of Sciences of the United States of America 84(24):8874-8878.
Owens, G. L., D. J. Rennison, W. T. Allison, and J. S. Taylor. 2012. In the four-eyed fish (Anableps anableps), the regions of the retina exposed to aquatic and aerial light do not express the same set of opsin genes. Biological Letters 8(1):86-89.
Owens, G. L., D. J. Windsor, J. Mui, and J. S. Taylor. 2009. A fish eye out of water: ten visual opsins in the four-eyed fish, Anableps anableps. PLoS One 4(6):e5970.
Palczewski, K., T. Kumasaka, T. Hori, C. A. Behnke, H. Motoshima, B. A. Fox, I. Le Trong, D. C. Teller, T. Okada, R. E. Stenkamp, M. Yamamoto, and M. Miyano. 2000. Crystal structure of rhodopsin: A G protein-coupled receptor. Science 289(5480):739-745.
Parry, J. W., K. L. Carleton, T. Spady, A. Carboo, D. M. Hunt, and J. K. Bowmaker. 2005. Mix and match color vision: tuning spectral sensitivity by differential opsin gene expression in Lake Malawi cichlids. Current Biology 15(19):1734-1739.
Parry, J. W. L., and J. K. Bowmaker. 2000. Visual pigment reconstitution in intact goldfish retina using synthetic retinaldehyde isomers. Vision Research 40(17):2241-2247.
Patzner, R. A., J. L. Van Tassell, M. Kovačić, and B. Kapoor. 2011. The biology of gobies. Science Publishers, Boca Raton, Florida.
Posada, D., and K. A. Crandall. 1998. MODELTEST: testing the model of DNA substitution. Bioinformatics 14(9):817-818.
Provencio, I., and R. G. Foster. 1993. Vitamin A 2-based photopigments within the pineal gland of a fully terrestrial vertebrate. Neuroscience Letters 155(2):223-226.
Saitou, N., and M. Nei. 1987. The neighbor-joining method: a new method for reconstructing phylogenetic trees. Molecular Biology and Evolution 4(4):406-425.
Shand, J., W. L. Davies, N. Thomas, L. Balmer, J. A. Cowing, M. Pointer, L. S. Carvalho, A. E. Trezise, S. P. Collin, L. D. Beazley, and D. M. Hunt. 2008. The influence of ontogeny and light environment on the expression of visual pigment opsins in the retina of the black bream, Acanthopagrus butcheri. Journal of Experimental Biology 211(Pt 9):1495-1503.
Shand, J., N. S. Hart, N. Thomas, and J. C. Partridge. 2002. Developmental changes in the cone visual pigments of black bream Acanthopagrus butcheri. Journal of Experimental Biology 205(Pt 23):3661-3667.
Sivasundar, A., and S. R. Palumbi. 2010. Parallel amino acid replacements in the rhodopsins of the rockfishes (Sebastes spp.) associated with shifts in habitat depth. Journal of Evolutionary Biology 23(6):1159-1169.
Spady, T. C., O. Seehausen, E. R. Loew, R. C. Jordan, T. D. Kocher, and K. L. Carleton. 2005. Adaptive molecular evolution in the opsin genes of rapidly speciating cichlid species. Molecular Biology and Evolution 22(6):1412-1422.
Takahashi, Y., and T. G. Ebrey. 2003. Molecular basis of spectral tuning in the newt short wavelength sensitive visual pigment. Biochemistry 42(20):6025-6034.
Takenaka, N., and S. Yokoyama. 2007. Mechanisms of spectral tuning in the RH2 pigments of Tokay gecko and American chameleon. Gene 399(1):26-32.
Tamura, K., and M. Nei. 1993. Estimation of the number of nucleotide substitutions in the control region of mitochondrial DNA in humans and chimpanzees. Molecular Biology and Evolution 10(3):512-526.
Tamura, K., G. Stecher, D. Peterson, A. Filipski, and S. Kumar. 2013. MEGA6: Molecular Evolutionary Genetics Analysis version 6.0. Molecular Biology and Evolution 30(12):2725-2729.
Terakita, A. 2005. The opsins. Genome Biology 6(3):213.
Ueno, Y., H. Ohba, Y. Yamazaki, F. Tokunaga, K. Narita, and T. Hariyama. 2005. Seasonal variation of chromophore composition in the eye of the Japanese dace, Tribolodon hakonensis. J Comp Physiol A Neuroethol Sens Neural Behav Physiol 191(12):1137-1142.
Wang, F. Y., W. C. Fu, I. L. Wang, H. Y. Yan, and T. Y. Wang. 2014. The giant mottled eel, Anguilla marmorata, uses blue-shifted rod photoreceptors during upstream migration. PLoS One 9(8):e103953.
Wang, F. Y., H. Y. Yan, J. S. Chen, T. Y. Wang, and D. Wang. 2009. Adaptation of visual spectra and opsin genes in seabreams. Vision Research 49(14):1860-1868.
Ward, M. N., A. M. Churcher, K. J. Dick, C. R. J. Laver, G. L. Owens, M. D. Polack, P. R. Ward, F. Breden, and J. S. Taylor. 2008. The molecular basis of color vision in colorful fish: Four Long Wave-Sensitive (LWS) opsins in guppies (Poecilia reticulata) are defined by amino acid substitutions at key functional sites. BMC Evolutionary Biology 8(1):210.
Yang, Z. 2007. PAML 4: phylogenetic analysis by maximum likelihood. Molecular Biology and Evolution 24(8):1586-1591.
Yang, Z., and J. P. Bielawski. 2000. Statistical methods for detecting molecular adaptation. Trends in Ecology & Evolution 15(12):496-503.
Yang, Z. H., and R. Nielsen. 2002. Codon-substitution models for detecting molecular adaptation at individual sites along specific lineages. Molecular Biology and Evolution 19(6):908-917.
Yang, Z. H., R. Nielsen, N. Goldman, and A. M. K. Pedersen. 2000. Codon-substitution models for heterogeneous selection pressure at amino acid sites. Genetics 155(1):431-449.
Yokoyama, S. 2000. Molecular evolution of vertebrate visual pigments. Progress in Retinal and Eye Research 19(4):385-419.
Yokoyama, S. 2008. Evolution of dim-light and color vision pigments. Annual Review of Genomics and Human Genetics 9:259-282.
Yokoyama, S., and B. Radlwimmer. 1998. The "five-sites" rule and the evolution of red and green color vision in mammals. Molecular Biology and Evolution 15(5):560-567.
Yokoyama, S., and T. Tada. 2003. The spectral tuning in the short wavelength-sensitive type 2 pigments. Gene 306:91-98.
Yokoyama, S., N. Takenaka, and N. Blow. 2007. A novel spectral tuning in the short wavelength-sensitive (SWS1 and SWS2) pigments of bluefin killifish (Lucania goodei). Gene 396(1):196-202.
Yokoyama, S., and R. Yokoyama. 1996. Adaptive evolution of photoreceptors and visual pigments in vertebrates. Annual Review of Ecology and Systematics 27:543-567.
Yokoyama, S., H. Zhang, F. B. Radlwimmer, and N. S. Blow. 1999. Adaptive evolution of color vision of the Comoran coelacanth (Latimeria chalumnae). Proceedings of the National Academy of Sciences of the United States of America 96(11):6279-6284.
Zhang, H., K. Futami, N. Horie, A. Okamura, T. Utoh, N. Mikawa, Y. Yamada, S. Tanaka, and N. Okamoto. 2000. Molecular cloning of fresh water and deep-sea rod opsin genes from Japanese eel Anguilla japonica and expressional analyses during sexual maturation. FEBS Letters 469(1):39-43.
Zhang, J. Z. 2003. Evolution by gene duplication: an update. Trends in Ecology & Evolution 18(6):292-298.