本實驗室研究孵化後10天吳郭魚腦部基因的表現，並建立吳郭魚腦部表現序列標記（expressed sequence tag, EST）資料庫。從中取得細胞增生相關功能性基因序列建構質體進行本論文之研究。使用細胞為老鼠神經瘤細胞（Neuro-2a）並以噻唑藍（3-(4,5)-dimethylthiahiazo (-z-y1)-3,5-di- phenytetrazoliumromide, MTT）進行細胞增生之分析。為探討實驗室已建構於Neuro-2a之穩定表現基因細胞株與細胞週期之關係，以同步化方式進行細胞週期同步化並以流式細胞儀進行研究進一步探討基因之表現對細胞週期之影響。實驗結果顯示Neuro-2a-CIRP穩定表現型細胞與Neuro-2a細胞株相比在細胞週期G2/M期停留時間有所延長。

Expressed sequence tags ( EST ) derived from the developing tilapia brain was cloned in our lab. In this study, the Neuro-2a cells was used to investigate the cell cycle –related gene. A stable cell line of Neuro-2a which was transfected with a cell cycel-related gene was established. The flow cytometer was used for the analysis of cell cycle. In a Neruo-2a-CIRP (cold-inducible RNA-binding protein,CIRP) stable cell line, the G2/M phase was prologate in cell cycle.

中文摘要---------------------------------------------------------------------- 1
英文摘要---------------------------------------------------------------------- 2
目錄---------------------------------------------------------------------------- 3
附圖、表目錄---------------------------------------------------------------- 4
前言---------------------------------------------------------------------------- 5
實驗材料與方法------------------------------------------------------------- 8
結果---------------------------------------------------------------------------- 21
討論---------------------------------------------------------------------------- 25
參考文獻---------------------------------------------------------------------- 30
附圖、表---------------------------------------------------------------------- 32

Aloia, R.C., J.K. Raison., (1989) Membrane function in mammalian hibernation. Biochim. Biophys. Acta. 988:123-146.
Burdon, R.H., (1987) Temperature and animal cell protein synthesis. Symp. Soc. Exp. Biol. 41:113-133.
Boonstra J., (2003) Progression through the G1-phase of the on-going cell cycle. J Cell Biochem. 90(2):244-52.
Chou R.C., Langan T.J., (2003) In vitro synchronization of mammalian astrocytic cultures by serum deprivation. Brain Res Brain Res Protoc. 2003 Jul;11(3):162-7.
Frisen J., Johansson C.B., Lothian C., Lendahl U. (1998) Central nervous system stem cells in the embryo and adult. Cell. Mol. Life Sci. 54(9):935-45.
Gasnereau I., Ganier O., Bourgain F., de Gramont A., Gendron M.C., Sobczak-Thépot J., (2007) Flow cytometry to sort mammalian cells in cytokinesis. Cytometry A. 2007 Jan;71(1):1-7
Ghosh A., Greenberg M.E., (1995) Distinct roles for bFGF and NT-3 in the regulation of cortical neurogenesis. Neuron 15: 89-103.
Hamid A.A., M. Mandai, et al., (2003) Expression of cold-inducible RNA-binding protein in the normal endometrium, endometrial hyperplasia, and endometrial carcinoma. Int J Gynecol Pathol 22(3): 240-7.
Jones P.G., M. Inouye, (1994) The cold-shock response-a hot topic. Mol. Microbiol. 11:811-818.
Liliensiek S.J., Schell K., Howard E., Nealey P., Murphy C.J., (2006) Cell sorting but not serum starvation is effective for SV40 human corneal epithelial cell cycle synchronization. Exp Eye Res. 2006 Jul;83(1):61-8. Epub 2006 Mar 20. Erratum in: Exp Eye Res. 2006 Dec;83(6):1521.
Morisaki T., Hirota T., Iida S., Marumoto T., Hara T., Nishiyama Y., Kawasuzi M., Hiraoka T., Mimori T., Araki N., Izawa I., Inagaki M., Saya H., (2002) WARTS tumor suppressor is phosphorylated by Cdc2/cyclin B at spindle poles during mitosis. FEBS Lett. Oct 9;529(2-3):319-24.
Nelson R. J., J. Kruuv, C.J. Koch, H.E. Fery, (1971) Effect of sub-optimal temperatures on survival of mammalian cells. Exp. Cell Res. 68:247-252.
Nishimune Y., T. Komatsu, (1972) Temperature-sensitiviyu of mouse testicular DNA synthesis in vitro. Exp. Cell Res. 75:514-517
Nishiyama H, Itoh K, Kaneko Y, Kishishita M, Yoshida O, Fujita J., (1997) A glycine-rich RNA-binding protein mediating cold-inducible suppression of mammalian cell growth. J Cell Biol. 19;137(4):899-908..
Oram S.W., Liu X.X., Lee T.L., Chan W.Y., Lau Y.F., (2006) TSPY potentiates cell proliferation and tumorigenesis by promoting cell cycle progression in HeLa and NIH3T3 cells. BMC Cancer. 9;6:154.
Rao P.N., J. Engelberg. (1965) HeLa cells: effects of temperature on the life cycle. Science 148:1092-1094.
Rieder C.L., Cole R.W., (2002) Cold-shock and the Mammalian cell cycle. Cell Cycle. 1(3):169-75..
Schorl C., Sedivy J.M., (2007) Analysis of cell cycle phases and progression in cultured mammalian cells. Methods. 41(2):143-50.
Sisken J.E., L. Morasca, S. Kibby, (1965) Effects of temperature on the kinetics of the mitotic cycle of mammalian cells in culture. Exp. Cell Res. 39:103-106.
Tsai C.L., L.H. Wang, Y.L. Shiue, T.Y. Chao (2007) Influence of temperature on the ontogenetic expression of neural development-related genes from developing Tilapia brain expressed sequence tags. Mar Biotechnol (NY). 9(2):243-61.
Urbani L., Sherwood S.W., Schimke R.T., (1995) Dissociation of nuclear and cytoplasmic cell cycle progression by drugs employed in cell synchronization. Exp Cell Res. 219(1):159-68.
Watanbe I., S. Okada, (1967) Stationary phase of cultured mammalian cells (L5178Y). J. Cell Biol. 35:285-294.
Whitfield M.L., Sherlock G., Saldanha A.J., Murray J.I., Ball C.A., Alexander K.E., Matese J.C., Perou C.M., Hurt M.M., Brown P.O., Botstein D.l., (2002) Identification of genes periodically expressed in the human cell cycle and their expression in tumors. Mol Biol Cell. 13(6):1977-2000.
Zhang E., Li X., Zhang S., Chen L., Zheng X., (2005) Cell cycle synchronization of embryonic stem cells: effect of serum deprivation on the differentiation of embryonic bodies in vitro. Biochem Biophys Res Commun. 2005 Aug 12;333(4):1171-7.