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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.author | I-Cheng Chen | en |
| dc.contributor.author | 陳怡辰 | zh_TW |
| dc.date.accessioned | 2021-07-01T08:13:00Z | - |
| dc.date.available | 2021-07-01T08:13:00Z | - |
| dc.date.issued | 2003 | |
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A research tool for studying cellular networks of protein interactions. Nucleic Acids Res. 30: 303-5. Xu, L., Weiner, B. M., and Kleckner, N. (1997) Meiotic cells monitor the status of the inerhomolog recombination complex. Genes Dev. 11: 106-118. Zakeri, Z. F., Wolgemuth, D. J., and Hunt, C. R. (1988) Identification and sequence analysis of a new member of the mouse HSP70 gene family and characterization of its unique cellular and developmental pattern of expression in the male germ line. Mol. Cell. Biol. 8: 2925-2932. Zhu, D., Dix, D. J., and Eddy, E. M. (1997) HSP70-2 is required for CDC2 kinase activity in meiosis I of mouse spermatocytes. Development 124: 3007-3014. | |
| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/75398 | - |
| dc.description.abstract | 老鼠熱休克蛋白質HSP70-2（70-kDa heat shock protein, HSP70）已知在減數分裂過程中扮演重要的角色，而酵母菌（budding yeast, Saccharomyces cerevisiae）中和老鼠HSP70-2蛋白質最為相似的是Ssa3（stress-seventy subfamily A）蛋白質，但其確切的功能目前還不清楚。本論文主要的目標在於研究酵母菌Ssa3蛋白質於減數分裂時期的功能。 首先，我們利用分子遺傳學方法獲得一些SSA3基因完全破壞的突變株（null mutants），結果發現ssa3突變本身對正常細胞的減數分裂沒有明顯的效應，但卻對zip1突變株的缺失檢控有影響。Zip1是酵母菌聯會複合體（synaptonemal complex, SC）中央區域的重要結構蛋白，在zip1突變株中，同源染色體無法聯會，細胞週期會停止（arrest）在粗絲期（pachytene）無法繼續完成減數分裂產生孢子。若在zip1突變株中將SSA3基因破壞，卻可以隱抑（suppress）zip1的缺失，使zip1突變株略過（bypass）粗絲期檢控點繼續進行減數分裂，但zip1ssa3的孢子存活率明顯比野生型低，顯示ssa3隱抑zip1不是透過修補zip1的缺失，而是ssa3在zip1中會影響粗絲期檢控點的作用。而ssa3突變無法使其他停止在粗絲期的突變株，如hop2或dmc1產生孢子。 利用蛋白質免疫轉印技術（Western blot analysis）分析Ssa3蛋白質，我們觀察到Ssa3蛋白質不只在減數分裂中才表現，但其表現量會隨著減數分裂的進行而增加。免疫螢光分析的結果顯示，不論是野生型或是zip1突變株，Ssa3蛋白質都同時存在於細胞核以及細胞質中；更進一步的觀察減數分裂粗絲期的染色體，發現Ssa3蛋白質位於野生型及zip1突變株的粗絲期染色體上，並且呈不完全連續的分佈。綜合以上的結果，我們推論在酵母菌進行減數分裂的過程中，Ssa3蛋白質可能參與維持染色體的適當構造（chromosome context），以使粗絲期檢控點能正確的作用。 | zh_TW |
| dc.description.abstract | The SSA3 (stress-seventy subfamily A) gene of Saccharomyces cerevisiae is a member of the HSP70 multigene family. To investigate the role of Ssa3 in meiosis, null mutants of SSA3 were generated and analyzed. ssa3 mutants undergo nearly wild-type levels of nuclear division and spore formation. Thus, Ssa3 is not required for sporulation in wild-type cells. However, ssa3 mutations can partially suppress a pachytene-arrest mutant, zip1, which has defects in chromosome structure and recombination. The spore viability of zip1ssa3 is about 41.6% of that of wild-type, suggesting that the recombination defects of zip1 persist in zip1ssa3 double mutants. These results indicate that ssa3 mutation might affect the normal function of the pachytene checkpoint in zip1 cells. Unlike zip1ssa3, ssa3 mutations fail to bypass other two pachytene-arrest mutants, hop2 and dmc1. Using epitope tagging strategy, the Ssa3 protein was detected by anti-HA antibodies. The Ssa3 protein is not a meiosis-specific protein, but its expression is slightly increased in meiosis. Chromosome spreading analyses indicate that Ssa3 protein is localized to pachytene chromosomes in both wild-type and the zip1 mutants. According to these results, we suggest that the Ssa3 protein may be involved in maintaining a proper chromosome structure for pachytene checkpoint signaling in budding yeast. | en |
| dc.description.provenance | Made available in DSpace on 2021-07-01T08:13:00Z (GMT). No. of bitstreams: 0 Previous issue date: 2003 | en |
| dc.description.tableofcontents | 中文摘要…………………………………………………………………i 英文摘要…………………………………………………………………ii 目錄………………………………………………………………………iii 表目錄……………………………………………………………………vi 圖目錄……………………………………………………………………vii 第一章 前言………………………………………………………………1 一、減數分裂簡介………………………………………………………1 二、減數分裂週期調控…………………………………………………2 三、老鼠HSP70-2蛋白質在減數分裂中之功用…………………………4 四、酵母菌Ssa3蛋白質…………………………………………………5 五、研究目標……………………………………………………………7 第二章 材料與方法………………………………………………………8 一、酵母菌品系及培養…………………………………………………8 二、一般性遺傳操作……………………………………………………8 1．細菌的轉型……………………………………………………………8 1．1 大腸桿菌勝任細胞的製備…………………………………………8 1．2 大腸桿菌的轉型……………………………………………………9 2．酵母菌的轉型…………………………………………………………9 3．孢子存活率分析……………………………………………………10 三、DNA製備……………………………………………………………10 1．一般性操作…………………………………………………………10 1．1 全細胞聚合?連鎖反應…………………………………………10 1．2 DNA電泳……………………………………………………………11 1．3 DNA片段純化………………………………………………………11 1．4 黏接反應…………………………………………………………12 1．5 質體DNA抽取………………………………………………………12 2．質體之建構…………………………………………………………13 四、酵母菌品系建構……………………………………………………14 1．酵母菌突變株之建構………………………………………………14 2．Ssa3-HA抗原決定基標定……………………………………………14 五、蛋白質分析…………………………………………………………15 1．蛋白質萃取…………………………………………………………15 2．蛋白質定量…………………………………………………………15 3．蛋白質電泳分析……………………………………………………16 4．蛋白質免疫轉印分析………………………………………………16 六、細胞學分析…………………………………………………………17 1．細胞核分裂分析……………………………………………………17 2．細胞免疫螢光定位…………………………………………………18 3．染色體螢光免疫分析………………………………………………19 4．影像分析……………………………………………………………20 第三章 結果……………………………………………………………21 一、ssa3突變株分析……………………………………………………21 1．ssa3突變對減數分裂無明顯的效應………………………………21 2．ssa3突變可以隱抑zip1突變株……………………………………22 3．ssa3突變無法隱抑其他停止在粗絲期的突變株…………………23 二、Ssa3蛋白質在減數分裂時期的表現………………………………25 1．Ssa3蛋白質表現量隨著進入減數分裂而增加……………………25 2．Ssa3蛋白質位於粗絲期染色體……………………………………25 第四章 討論……………………………………………………………27 一、酵母菌zip1突變株之隱抑子………………………………………27 二、染色體蛋白質與粗絲期檢控點之關聯……………………………28 三、ssa3隱抑zip1突變株之可能機制…………………………………31 四、其他可能參與減數分裂之熱休克蛋白質…………………………32 參考文獻…………………………………………………………………33 圖表………………………………………………………………………40 附錄………………………………………………………………………59 | |
| dc.language.iso | zh-TW | |
| dc.title | 酵母菌Ssa3蛋白質於減數分裂時期功能之研究 | zh_TW |
| dc.title | The function of yeast Ssa3 protein in meiosis | en |
| dc.date.schoolyear | 91-2 | |
| dc.description.degree | 碩士 | |
| dc.relation.page | 59 | |
| dc.rights.note | 未授權 | |
| dc.contributor.author-dept | 生命科學院 | zh_TW |
| dc.contributor.author-dept | 植物科學研究所 | zh_TW |
| 顯示於系所單位： | 植物科學研究所 | |
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