https://scholars.lib.ntu.edu.tw/handle/123456789/194420
標題: | GTP CYCLOHYDROLASE I 基因轉殖小鼠啟動子表現及表現型分析 | 作者: | 胡務亮 | 關鍵字: | GTP cyclohydrolase I;Dopa-敏感性肌肉張力不全症;基因轉殖小鼠;Dopa-responsive dystonia;transgenic mice | 公開日期: | 2001 | 出版社: | 臺北市:國立臺灣大學醫學院小兒科 | 摘要: | GTP cyclohydrolase I (GCH)負責 tetrahydrobiopterine(BH4)合成的第一個步驟。BH4 是一個很重要的分子,它是phenylalanine hydroxylase, tyrosine hydroxylase, tryptophan hydroxylase以及nitric oxide synthase的輔因子。GCH 基因突變會引起惡性苯酮尿症或Dopa-敏感性肌肉 張力不全症,兩者和巴金斯症一樣,都需要接受 L-dopa治療。大部分Dopa-敏感性肌肉張力不全症 是顯性遺傳,其分子機轉為dominant-negative。基 於BH4生理作用的廣泛性,其引發疾病之複雜性, 以及與巴金斯症的相關性,我們希望能製造出缺乏 GCH的動物模型。 我們在過去的時間中,我們自人類基因庫中 篩選到GCH基因前方一斷13kb長的序列。並且證實 2.8kb及5kb(包含部分exon 1)的片段可以表現出啟 動子活性。我們將2.8kb片段後方接上LacZ基因後 注入小鼠受精卵中,結果LacZ F1小鼠腦部無法見 到LacZ基因的表現。因此我們將含有Dopa-敏感性 肌肉張力不全症G201E突變的GCH基因cDNA,接 在5kb片段後方。小鼠F0及F1均已篩選完畢。 在F0及F1的轉殖小鼠中,我們發現有一些小 鼠在Rota Rod中的表現不佳。這些小鼠有可能是腦 中GCH基因的表現受到影響,所以表現出運動上的 障礙。然而這些小鼠的子代並沒有辦法穩定的表現 出這樣的異常,所以無法證實運動障礙和轉殖基因 間的關係。 當轉殖小鼠年齡增大後,我們發現其中有一 些有異常的動作。這是一種持續性的沒有目的的旋 轉。來自同一祖先的好幾株都可以看到這樣的異 常。這可能是一種晚發性的運動障礙,很可能和轉 殖基因,也就是突變型的GCH基因有關。這些小鼠 的生殖力差,新生小鼠死亡率高,相關組織學分析 正在進行中。 GTP cyclohydrolase I (GCH) is responsible for the first step of tetrahydrobiopterin (BH4) biosynthesis. BH4 is an important molecule, because it is the cofactor of phenylalanine hydroxylase, tyrosine hydroxylase, tryptophan hydroxylase and nitric oxide synthase. GCH gene mutations are associated with wither malignant phenylketonuria and dopa-responsive dystonia (DRD). Both of them are responsive to L-dopa treatment. DRD is mostly dominantly inherited, and through the dominant-negative mechanism. In the past years, we have isolated a 13kb DNA from human genomic library containing the 5’ region (including part of exon 1) of GCH gene. We have shown that both a 2.8 and 5kb fragments possessed promoter activity. We connected the 2.8kb fragment to LacZ gene. The construct was injected into mouse fertilized egg. We have screened F0 and F1 for the LacZ mice, but the gene was not expressed. We inserted a GCH cDNA containing the G201E DRD mutation after the 5kb promoter. Both F0 and F1 G201E mice were produced. The phenotype of the transgenic mice was checked by Rota-Rod. Several mice were found to have poor performance on the Rota. These mice may have decreased GCH activity because of the expression of the dominant negative GCH gene. However, the offspring of these mice could not express stably the same phenotype. Therefore, we can not make conclusion on the association between motor defect and the transgene. When the mice were getting old, some of them demonstrated behavior problem. They tended to make purposeless circling movement. This could be a late onset motor abnormality, and may be associated with the GCH gene. Some mice were from the same ancestor. These mice have low fertility power, and many of their babies died shortly after birth. Further studies on them are going now. |
URI: | http://ntur.lib.ntu.edu.tw//handle/246246/22855 | 其他識別: | 892314B002387 | Rights: | 國立臺灣大學醫學院小兒科 |
顯示於: | 醫學系 |
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892314B002387.pdf | 286.7 kB | Adobe PDF | 檢視/開啟 |
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