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研究生:史承治
研究生(外文):Cheng-chih Shih
論文名稱:脂肪細胞受各種荷爾蒙調控基因之選殖與性質分析
論文名稱(外文):Cloning and characterization of genes that are responsive to various hormones in 3T3-L1 adipocytes
指導教授:馮濟敏
指導教授(外文):Jim C. Fong
學位類別:碩士
校院名稱:國立陽明大學
系所名稱:生物化學研究所
學門:生命科學學門
學類:生物化學學類
論文種類:學術論文
論文出版年:2004
畢業學年度:92
語文別:中文
論文頁數:70
中文關鍵詞:脂肪細胞荷爾蒙調控之基因
外文關鍵詞:3T3-L1 adipocytesCloning and characterization of genes that are responsive to various hormones
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中文摘要
本實驗室利用3T3-L1 脂肪細胞當作實驗材料去研究一些新基因的特性。在加入不同的藥物處理後,我們利用suppression subtractive hybridization (SSH)的方法產生一些subtractive cDNA clones。接著我們利用生物資訊的方法來分析這些clones。在基因的篩選過程中,我們針對兩個目前功能未知的基因作為研究目標。初步的實驗結果顯示,這兩個基因的產物都具有不同的splicing form。本篇論文即在探討此兩個基因的性質及其蛋白質可能的功能。
第一個基因在NCBI 資料庫中的accession number 為AK077556 (long form)。利用生物資訊的方法發現,這個基因具有另外兩個不同的splicing form其accession number分別為:short form (BC012275),和middle form (AK037644)。從蛋白質的資料庫中得知long form及middle form都具有zf-MYND (conserved zinc finger- hydrophobic heptad repeat) 和ANK (ankyrin repeats) domains。short form則只有ANK domain。由RT-PCR的實驗結果發現,此三種splicing form對於各種不同的荷爾蒙刺激有不同的反應。
第二個基因的accession number 為BC009125,從蛋白質的資料庫中得知其具有modified RING finger domain。此基因splicing form的發現則在於PCR的洋菜膠電泳實驗中。在每次的電泳實驗中除了target band (744bp)外,都會有另一個minor band (592bp)。由RT-PCR的實驗結果發現,此兩種splicing form對於各種不同的荷爾蒙刺激也有不同的反應。
在資料庫中的比對發現,這兩基因在其它物種之間也具有同源性的基因,包括人類在內。實驗結果發現,這兩個基因也存在於老鼠各種不同的組織及人類的一些癌症細胞株中。
至於這兩個基因的功能為何?是否與癌症的生長有關,還需要進一步的實驗去求證。
Abstract
Our lab used 3T3-L1 adipocyte as materials to study some novel genes. Ather treating cells with various substances, suppression subtractive hybridization ( SSH) was employed to generate some subtractive cDNA clones. These EST clones were analyzed by bioinformatics. In the progress of finding some novel genes, we chose two genes as our study target, and the functions of the two genes are unknown so far. But analyzing the databases, we found that the products of these two genes have different splicing forms. The purpose of this thesis is to explore the properties and functions of these two genes.
In the NCBI database, the accession number of the first gene is AK077556 (long form). We made use of bioinformatics to search some information about this gene, and we got that this gene has the other two splicing forms. The accession number of these two splicing forms is BC012275 (short form) and AK037644 (middle form), respectively. From the protein database, we knew that long form and middle form both have zf-MYND (conserved zinc finger- hydrophobic heptad repeat) and ANK (ankyrin repeats) domains, but short form only has ANK domain. From the result of RT-PCR (Reverse transcription – Polymerase Chain Reaction ), we found that these three splicing forms have different reaction patterns treating with varios substances.
The accession number of the second gene is BC009125. Analyzing the sequence of this protein, we knew that the protein has U box (modified RING finger) domain. But the other splicing form of this gene is out of our expectation. Because we didn’t find it in the alternative splicing database. Interestingly, we found it from agarose gel electrophoresis. There always has another band under our target band. From the result of RT-PCR, we found that these two splicing forms also have different patterns treating with various substances.
By analyzing the information in databases, we noted that these two genes have homology among species, including homo sapiens. We also found that these two genes maybe involved in some cancer cell lines. So, there is still a long way to go to figure out the functions of these two genes.
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