現在位置首頁 > 博碩士論文 > 詳目
  • 同意授權
論文中文名稱:參與鏈黴菌SCP1質體端粒複製的Tac及Tpc蛋白 [以論文名稱查詢館藏系統]
論文英文名稱:The functions of Tac and Tpc in the terminal patching of Streptomyces SCP1 plasmid [以論文名稱查詢館藏系統]
院校名稱:臺北科技大學
學院名稱:工程學院
系所名稱:生物科技研究所
中文姓名:簡佑年
英文姓名:Yu-Nien Chien
研究生學號:94688013
學位類別:碩士
語文別:中文
口試日期:2007-07-26
論文頁數:61
指導教授中文名:黃志宏
指導教授英文名:Chih-Hung Huang
口試委員中文名:陳文盛;楊千金
口試委員英文名:Carton W. Chen;Chien-Chin Yang
中文關鍵詞:鏈黴菌末端蛋白線型質體SCP1
英文關鍵詞:StreptomycesSCP1telomereTacTpcterminal protein
論文中文摘要:線形染色體和線形質體在鏈黴菌中是常見的。這些線形的複製子有幾個共同的特徵,例如具有許多廻文序列和5’端上共價鍵結著末端蛋白(TPs)。在大多數鏈黴菌的染色體和質體,末端蛋白的結構及端粒DNA序列非常相似。大多數的端粒DNA序列彼此相似,其末端蛋白彼此的蛋白質序列也相似。發現在許多末端蛋白的基因上游,有一個和末端修復相關蛋白的基因tap也很相似。在DNA複製時,Tap會和pSLA2的末端序列以及末端蛋白結合,因此,Tap在末端複製時帶著末端蛋白到末端序列進行末端的修復。少數的鏈黴菌質體如pRL1、pRL2 (Qin et al., 2006)及SCP1 (Kinashi et al.,1991),以及S. griseus的染色體,這些的端粒序列及末端蛋白和大多數不相似,目前研究較清楚的如SCP1,SCP1的端粒相關蛋白(Tac)基因也座落在末端蛋白(Tpc)基因的上游,由於這兩套端粒系統有些特徵相似,因此也可能以相同的機制進行末端的修復。為了探究這個問題,首先我們以酵母菌雙雜交法去測試Tac和Tpc的交互作用,發現這兩個蛋白在酵母菌中沒有交互作用。其次,在SCP1上tac、SCP1.126和tpc三個基因相當靠近,我們將這個卡匣中含有tpc的DNA片段反向,發現無法支持SCP1的末端修復。最後為了後續的生物細胞研究和蛋白質的純化,我們在Tac和Tpc的C端和N端分別加上組胺酸標記,初步結果顯示帶有組胺酸標記的Tac和Tpc在鏈黴菌裡可能失去其活性,於是將再測試標定其他種標記。
論文英文摘要:The linear chromosomes and plasmids are common in Streptomyces. The telomeres of these linear replicons share the same characters, like palindrome rich sequences and 5’capped terminal proteins (TP). In Streptomyces, the TPs and telomeric DNA can be separate to two pupolations by their sequences and structures. In the first population, their telomeric DNA are similar to each other, and their capped TPs are similar to each other, too. A terminal patching required gene, tap, is found on the upstream of several TP genes of the first group. The Tap binds the replicating telomeric DNA of pSLA2 and TP, therefore, Tap play a role to recruit TP to replicating telomeres for further terminal patching. In the second population, their telomeric DNA sequences can’t align with those in first population. The best studied telomere of this population is SCP1, with the unique telomeric DNA and capped TP sequences. Its tpc (the gene of TP of SCP1) is located in SCP1, and a terminal patching required gene, tac, is on the upstream of tpc. Because some characters are shared between these two telomere systems, they may proceed with their terminal patching by the same mechanism. For addressing this question, we firstly tested the interaction of Tac and Tpc by yeast two-hybridize system and found they didn’t interact in yeast. Secondly, the tac, SCP1.126 and tpc are close together on SCP1, we reversed the direction of a DNA fragment containing tpc in this DNA cassette, and found this failed to support terminal patching of SCP1. This preliminary result hints that three ORFs may locat on an operon. Finally, we were added his-tag on the C-terminal and N-terminal of Tac and Tpc for further cell biology study and protein purification. The preliminary result showed these his-tag fused Tac and Tpc lost their activity in vivo.The other tag lablled strategy will be test soon.
論文目次:中文摘要 i
英文摘要 iii
誌謝 v
目錄 vi
圖目錄 vii
第一章 序論 1
第二章 材料與方法 5
第三章 結果 9
第四章 討論 38
參考文獻 41
附錄 44
論文參考文獻:[1] J. Sambrook and D.W. Russell, Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press, 2001.
[2] T. Kieser, M. J. Chater et al, Practical Streptomyces genetics, The John Innes Foundation, 2000.
[3] T.A. Brown edited, Molecular Biology Labfax, Blackwell scientific publications, 1991.
[4] Bao, K. and S. N. Cohen (2001). "Terminal proteins essential for the replication of linear plasmids and chromosomes in Streptomyces." Genes Dev 15(12): 1518-27.
[5] Bao, K. and S. N. Cohen (2003). "Recruitment of terminal protein to the ends of Streptomyces linear plasmids and chromosomes by a novel telomere-binding protein essential for linear DNA replication." Genes Dev 17(6): 774-85.
[6] Bentley, S. D., S. Brown, et al. (2004). "SCP1, a 356,023 bp linear plasmid adapted to the ecology and developmental biology of its host, Streptomyces coelicolor A3(2)." Mol Microbiol 51(6): 1615-28.
[7] Blackburn, E. H. (2001). "Switching and signaling at the telomere." Cell 106(6): 661-73.
[8] Calcutt, M. J. and F. J. Schmidt (1992). "Conserved gene arrangement in the origin region of the Streptomyces coelicolor chromosome." J Bacteriol 174(10): 3220-6.
[9] Chang, P. C. and S. N. Cohen (1994). "Bidirectional replication from an internal origin in a linear streptomyces plasmid." Science 265(5174): 952-4.
[10] Emre, N. C. and S. L. Berger (2006). "Histone post-translational modifications regulate transcription and silent chromatin in Saccharomyces cerevisiae." Ernst Schering Res Found Workshop(57): 127-53.
[11] Gao, L. and D. S. Gross (2006). "Using genomics and proteomics to investigate mechanisms of transcriptional silencing in Saccharomyces cerevisiae." Brief Funct Genomic Proteomic 5(4): 280-8.
[12] Greider, C. W. and E. H. Blackburn (1996). "Telomeres, telomerase and cancer." Sci Am 274(2): 92-7.
[13] Hirochika, H. and K. Sakaguchi (1982). "Analysis of linear plasmids isolated from Streptomyces: association of protein with the ends of the plasmid DNA." Plasmid 7(1): 59-65.
[14] Hopwood, D. A., T. Kieser, et al. (1983). "Plasmids, recombination and chromosome mapping in Streptomyces lividans 66." J Gen Microbiol 129(7): 2257-69.
[15] Huang, C. H., Y. S. Lin, et al. (1998). "The telomeres of Streptomyces chromosomes contain conserved palindromic sequences with potential to form complex secondary structures." Mol Microbiol 28(5): 905-16.
[16] Huang, C. H., H. H. Tsai, et al. (2007). "The telomere system of the Streptomyces linear plasmid SCP1 represents a novel class." Mol Microbiol 63(6): 1710-8.
[17] Kinashi, H., M. Shimaji-Murayama, et al. (1991). "Nucleotide sequence analysis of the unusually long terminal inverted repeats of a giant linear plasmid, SCP1." Plasmid 26(2): 123-30.
[18] Lin, Y. S., H. M. Kieser, et al. (1993). "The chromosomal DNA of Streptomyces lividans 66 is linear." Mol Microbiol 10(5): 923-33.
[19] Musialowski, M. S., F. Flett, et al. (1994). "Functional evidence that the principal DNA replication origin of the Streptomyces coelicolor chromosome is close to the dnaA-gyrB region." J Bacteriol 176(16): 5123-5.
[20] Qin, Z. and S. N. Cohen (1998). "Replication at the telomeres of the Streptomyces linear plasmid pSLA2." Mol Microbiol 28(5): 893-903.
[21] Shiffman, D. and S. N. Cohen (1992). "Reconstruction of a Streptomyces linear replicon from separately cloned DNA fragments: existence of a cryptic origin of circular replication within the linear plasmid." Proc Natl Acad Sci U S A 89(13): 6129-33.
[22] Takano, E., J. White, et al. (1995). "Construction of thiostrepton-inducible, high-copy-number expression vectors for use in Streptomyces spp." Gene 166(1): 133-7.
[23] Yang, C. C., Y. H. Chen, et al. (2006). "In vitro deoxynucleotidylation of the terminal protein of Streptomyces linear chromosomes." Appl Environ Microbiol 72(12): 7959-61.
[24] Yang, C. C., C. H. Huang, et al. (2002). "The terminal proteins of linear Streptomyces chromosomes and plasmids: a novel class of replication priming proteins." Mol Microbiol 43(2): 297-305.
[25] Zakrzewska-Czerwinska, J. and H. Schrempf (1992). "Characterization of an autonomously replicating region from the Streptomyces lividans chromosome." J Bacteriol 174(8): 2688-93.
論文全文使用權限:同意授權於2008-08-27起公開