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  1. NTU Theses and Dissertations Repository
  2. 醫學院
  3. 醫學檢驗暨生物技術學系
請用此 Handle URI 來引用此文件: http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/38769
完整後設資料紀錄
DC 欄位值語言
dc.contributor.advisor方偉宏(Woei-horng Fang)
dc.contributor.authorWei-Tsan Chinen
dc.contributor.author秦維璨zh_TW
dc.date.accessioned2021-06-13T16:45:26Z-
dc.date.available2006-08-02
dc.date.copyright2005-08-02
dc.date.issued2005
dc.date.submitted2005-06-29
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Bill, C. A., Taghian, D. G., Duran, W. A., and Nickoloff, J. A. (2001) Repair bias of large loop mismatches during recombination in mammalian cells depends on loop length and structure. Mutat Res. 485, 255-265
Bollag, R. J., Elwood, D. R., Tobin, E. D., Godwin, A. R., and Liskay, R. M. (1992) Formation of heteroduplex DNA during mammalian intrachromosomal gene conversion. Mol Cell Biol. 12, 1546-1552
Burdett, V., Baitinger, C., Viswanathan, M., Lovett, S. T., Modrich, P. (2001) In vivo requirement for RecJ, ExoVII, ExoI, and ExoX in methyl-directed mismatch repair. Proc. Natl. Acad. Sci. U.S.A. 98, 6765-6770
Carraway, M., and Marinus, M. G. (1993) Repair of Heteroduplex DNA molecules with multibase loops in Escherichia coli. J. Bacteriol. 175, 3972-3980
Church, G. M., and Gilbert, W. (1984) Genomic sequencing. Proc. Natl. Acad. Sci. U. S. A. 81, 1991-1995
Corrette-Bennett, S. E., Parker, B.O., Mohlman N.L., and Lahue, R. S. (1999) Correction of large mispaired DNA loops by extracts of Saccharomyces cerevisiae. J. Biol. Chem. 274, 17605-17611
Corrette-Bennett, S. E., Mohlman, N. L., Rosado, Z., Miret, J. J., Hess, P. M., Parker, B. O., and Lahue, R. S. (2001) Efficient repair of large DNA loops in Saccharomyces cerevisiae. Nucleic Acids Res. 29, 4134-4143
Corrette-Bennett, S. E., Borgeson, C., Sommer, D., Burgers, P. M., and Lahue, R. S. (2004) DNA polymerase delta, RFC and PCNA are required for repair synthesis of large looped heteroduplexes in Saccharomyces cerevisiae. Nucleic Acids Res. 32, 6268-6275
Deng, W. P., and Nickoloff, J. A. (1994) Mismatch repair of heteroduplex DNA intermediates of extrachromosomal recombination in mammalian cells. Mol. Cell. Biol. 14, 400-406
Fang, W. H., and Modrich, P. (1993) Human strand-specific mismatch repair occurs by a bidirectional mechanism similar to that of the bacterial reaction. J. Biol. Chem. 268. 11838-11844
Fang, W. H., Wu, J. Y., and Su, M. J. (1997) Methyl-directed repair of mismatched small Heterologous sequences in cell extracts from Escherichia coli. J. Biol. Chem. 272, 22714-22720
Fang, W. H., Wang, B. J., Wang, C. H., Lee, S. J., Chang, Y. T., Chuang, Y. K., and Lee, C. N. (2003) DNA loop repair by Escherichia coli cell extracts. J. Biol. Chem. 278, 22446-22452
Genschel, J., Littman, S. J., Drummond, J. T., and Modrich, P. (1998) Isolation of MutSbeta from human cells and comparison of the mismatch repair specificities of MutSbeta and MutSalpha. J. Biol. Chem. 273, 19895-19901
Jiricny, J. (1998) Replication errors: cha(lle)nging the genome. EMBO J. 17, 6427-6436
Lahue, R. S., Su, S. S., and Modrich, P. (1987) Requirement for d(GATC) sequence in Escherichia coli mutHLS mismatch correction. Proc. Natl. Acad. Sci. U. S. A. 84, 1482-1486
Lahue, R. S., Au, K. G., and Modrich, P. (1989) DNA mismatch correction in a defined system. Science 245, 160-164
Littman, S. J., Fang, W. H., and Modrich, P. (1999) Repair of large insertion/deletion heterologies in human nuclear extracts is directed by a 5’ single-strand break and is independent of the mismatch repair system. J. Biol. Chem. 274, 7474-7481
Lu, A. L., Clark, S., and Modrich, P. (1983) Methyl-directed repair of DNA base-pair mismatches in vitro. Proc. Natl. Acad. Sci. U. S. A. 80, 4639-4643
Horst, J. P., Wu, T. H., and Marinus, M. G. (1999) Escherichia coli mutator genes. Trends in Microbiol. 7, 29-36
Miret, J. J., Parker, B. O., and Lahua, R. S., (1996) Recognition of DNA insertion/deletion mismatches by an activity in Saccharomyces cerevisiae. Nucleic Acids Res. 24. 721-9
McCulloch, S. D., Gu, L., and Li, G. M. (2003) Bi-directional processing of DNA loops by mismatch repair-dependent and -independent pathways in human cells. J. Biol. Chem. 278, 3891-3896
McCulloch, S. D., Gu, L., and Li, G. M. (2003) Nick-dependent and -independent processing of large DNA loops in human cells. J. Biol. Chem. 278, 50803-50809
Modrich, P., and Lahue, R. (1996) Mismatch repair in replication fidelity, genetic recombination and cancer biology. Annu. Rev. Biochem. 65, 101-133
Modrich, P. (1997) Strand-specific mismatch repair in mammalian cells. J. Biol. Chem. 272, 24727-24730
Nag, D. K., White, M. A., and Petes, T. D. (1989) Palindromic sequences in heteroduplex DNA inhibit mismatch repair in yeast. Nature 340, 318-320
Ohshima, K., and Wells, R. D. (1997) Hairpin formation during DNA synthesis primer realignment in vitro in triplet repeat sequences from human hereditary disease genes. J. Biol. Chem. 272, 16798-16806
Parker, B. O., and Marinus, M. G. (1992) Repair of DNA Heteroduplexes containing small Heterologous sequences in Escherichia coli. Proc. Natl. Acad. Sci U. S. A. 89, 1730-1734
Pearson, C. E., Ewel, A., Acharya, S., Fishel, R. A., and Sinden, R. R. (1997) Human MSH2 binds to trinucleotide repeat DNA structures associated with neurodegenerative diseases. Hum. Mol. Genet. 6, 1117-1123
Pearson, C. E., and Sinden, R. R. (1998) Trinucleotide repeat DNA structures: dynamic mutations from dynamic DNA. Curr. Opin. Struct. Biol. 8, 321-330
Radman, M., Matic, I., Halliday, J. A., and Taddei, F. (1995) Editing DNA replication and recombination by mismatch repair: from bacterial genetics to mechanisms of predisposition to cancer in humans. Philos. Trans. R. Soc. Lond. B. Biol. Sci. 347, 97-103
Sia, E. A., Kokoska, R. J., Dominska, M., Greenwell, P., and Petes, T. D. (1997) Microsatellite instability in yeast: dependence on repeat unit size and DNA mismatch repair genes. Mol. Cell. Biol. 17, 2851-2858
Su, S. S., Lahue, R. S., Au, K. G., and Modrich, P. (1988) Mispair specificity of methyl-directed DNA mismatch correction in vitro. J. Biol. Chem. 263, 6829-6835
Umar, A., Boyer, J. C., and Kunkel, T. A., (1994) DNA loop repair by human cell extracts. Nature 266, 814-816
Weng, Y. S., and Nickoloff, J. A. (1998) Evidence for independent mismatch repair processing on opposite sides of a double-strand break in Saccharomyces cerevisiae. Genetics 148, 59-70
Wierdl, M., Dominska, M., and Petes, T. D. (1997) Microsatellite instability in yeast: dependence on the length of the microsatellite. Genetics 146, 769-779
Viswanathan, M., and Lovett, S. T. (1999) Exonuclease X of Escherichia coli. J. Biol. Chem. 274, 30094-30100
Viswanathan, M., Burdett, V., Baitinger, C., Modrich, P., and Lovett, S. T. (2001) Redundant exonuclease involvement in Escherichia coli methyl-directed mismatch repair. J. Biol. Chem. 276, 31053-31058
李淑貞, (2002) 核酸環與鹼基配對錯誤在大腸桿菌萃取液中之共同修復,國立台灣大學碩士論文
張友婷, (2003) 大腸桿菌修復髮夾核酸配對錯誤之機制, 國立台灣大學碩士論文
莊以光, (2003) 人類細胞對髮夾結構之修復機制,國立台灣大學碩士論文
dc.identifier.urihttp://tdr.lib.ntu.edu.tw/jspui/handle/123456789/38769-
dc.description.abstract生物為了維持基因體的穩定,演化出許多不同機制來修復核酸上的錯誤,以避免個體產生病變,甚至死亡。大型核酸環修復系統是近年來才被發現的修復機制,到目前為止已知,在大腸桿菌中此修復機制有下列的特徵:修復過程是由斷股所指引(nick-directed)、需要鎂離子與dNTPs的參與、不需鹼基配對錯誤修復系統的蛋白MutHLS參與、不需要外加ATP來提供能量;此外本實驗室還發現,髮夾結構修復的特徵與大型核酸環相同,因此這兩種錯誤可能由相同機制來修復,但到目前為止,詳細的修復機制與所參與的蛋白卻依然不清楚。因此,為了找出參與修復反應的蛋白,我們取得8種不同核酸修復缺陷的大腸桿菌突變株,利用其細胞萃取液進行測試,結果發現異雙股核酸上的修復反應並沒有被抑制,因此可知UvrA、UvrB、UvrC、UvrD、RecJ、ExoI、ExoVII及SbcCD這8種蛋白,並沒有參與大型核酸環及髮夾結構的修復。在修復路徑 (repair patch)的探討方面,我們藉著不加入dNTPs,或加入ddNTPs,來限制修復反應的進行,並以鹼性瓊酯膠電泳 (alkaline agarose gel),對反應中所產生的中間產物進行初步分析,再利用變性聚丙醯胺凝膠電泳 (denaturing polyacrylamide gel),對信號所在區域做細部分析,結果發現大型核酸環與髮夾結構上發生特異性斷股,表示未配對序列可能經由斷股而被移除。了解切除反應 (excision)的範圍,我們利用限制酵素來分析斷股與未配對序列間的核酸是否被移除,結果發現未配對序列兩側的核酸,皆保持雙股狀態。綜合以上結果,我們推測在大腸桿菌中,大型核酸環與髮夾結構的修復,皆由斷股所指引,並直接於未配對序列上進行切割,再進行的修復,但其詳細機制則仍需更進一步的研究。zh_TW
dc.description.abstractDNA loop and hairpin are products of normal DNA biosysthetic errors or homeologous recombination and can lead to severe genomic instability if unrepaired. Previous studies showed that a strand break located either 3’ or 5’ to the large loop is sufficient to direct repair to the nicked strand in Escherichia coli cell extracts. This activity is distinct from mismatch repair pathway. Furthermore, our previous results suggested that hairpin and large loop structures would be processed by the same mechanism because of their similar repair characteristics. To investigate what components are involved and understand how heterologies are processed, we used a set of heteroduplexes containing an insertion/deletion large loop or hairpin for our study. Heteroduplexes were tested in the extracts from different E. coli mutant strains for repair efficiency. The results indicated that the correction of large loop and hairpin repair are independent of uvrA, uvrB, uvrC, uvrD, recJ, exoI, exoVII and sbcCD gene products. By limiting the repair synthesis in reactions, the incision or excision intermediates can be trapped and analyzed by denaturing gel electrophoresis. We found that strand- and loop-specific incisions are in the proximity to the heterologies. To further determine the involvement of excision reactions in large loop repair, we employed restriction enzyme digestion assay to the repair intermediates. Our data suggested that large loop and hairpin are processed by specific incision in the heterologies, not processed by the extensive excision from the pre-existing nick toward heterologies.en
dc.description.provenanceMade available in DSpace on 2021-06-13T16:45:26Z (GMT). No. of bitstreams: 1
ntu-94-R92424020-1.pdf: 1752195 bytes, checksum: 2cd3187f3e88731d638e5b94e8db1b39 (MD5)
Previous issue date: 2005
en
dc.description.tableofcontents中文摘要 1
英文摘要 2
縮寫表 3
前言 4
材料與方法 9
一、菌株 9
二、大腸桿菌細胞萃取物之製備 9
三、f1P突變噬菌體之建構 10
四、f1P系列雙股核酸之製備 11
五、f1P系列單股核酸之製 12
六、異雙股核酸之建構 13
七、試管中之修復反應 14
八、探針標定之方法 15
九、修復反應之中間產物分析 15
十、南方墨點法之步驟 16
結果 17
一、異雙股核酸的選擇 17
二、試管中異雙股核酸之修復分析 17
三、參與修復反應蛋白之分析 18
四、限制情況下之修復效率分析 19
五、修復反應中間產物之初步分析 20
六、修復反應中間產物之高解析分析 21
七、大型核酸環修復反應中核酸移除範圍之分析 23
討論 25
附圖 29
附表 42
參考文獻 44
dc.language.isozh-TW
dc.title大腸桿菌中大型核酸環修復路徑之分析zh_TW
dc.titleRepair Patch Analysis of Large Loop Repair in Escherichia colien
dc.typeThesis
dc.date.schoolyear93-2
dc.description.degree碩士
dc.contributor.oralexamcommittee許濤(Todd Hsu),蔡芷季(Jyy-Jin Tsai),王憶卿(Yi-Ching Wang)
dc.subject.keyword核酸環修復,大腸桿菌,zh_TW
dc.subject.keywordloop repair,E. coli,en
dc.relation.page48
dc.rights.note有償授權
dc.date.accepted2005-06-30
dc.contributor.author-college醫學院zh_TW
dc.contributor.author-dept醫事技術學研究所zh_TW
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