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標題: | 酵母菌分泌性載體之構築 Construction of Secretion Vectors in Saccharomyces cerevisiae |
作者: | Dz-Chi Chen 陳子智 |
出版年 : | 1986 |
學位: | 碩士 |
摘要: | 本論文之主旨乃在於構築—酵母菌分泌性載體系統,並經由字碼子讀框校正;試圖將B型肝炎前表面抗原基因(pre S2—SAg)經由此分泌性載體系統,在酵母菌之分泌途徑表現。 為建立酵母菌分泌載體系統,首先將-帶有酵母菌α因數(費洛蒙)之起動子—前導序列的載體pYA41上之多重選殖區,進行字碼子讀框校正,而創造出不同之讀框,以利外來基因在插入此多重選殖區後,仍保持正確之字碼子讀框,在此步驟中共得到pYA41′/35、pYA41′/45、pUR 222/α1、pUC9/α1、pUC9/α2、pUC19/α1、pUC19/α2等七種載體,各具有不同之讀框及不同多重選殖區之分泌性載體系統,可供黏端黏接或鈍端黏接,用來插入外來蛋白基因。 為便利此系統能在酵母菌轉形系統中使用,乃將這些分泌性載體與本實驗室原有之帶抗藥性及營養需求性標誌之酵母菌載體結合,而構築出YAUT0、YAUT1、YAUT2、pUC9/α1S、YAMTS、YAUTS、YAMKS七種具有遺傳標誌之分泌性載體。其中後四種已與B型肝炎病毒表面抗原基因結合,在黏接處之字碼子讀框已子校正,以期能在酵母菌中分泌出B型肝炎病毒表面抗原。 In this thesis, a yeast secretion vector system was developed. To investigate whether the secretion vector system is able to allow effective processing and secretion of heterologous protein in yeast, a hepatitis B virus gene coding for preS2SAg was introduced into the down stream of yeast α-factor-promoter-leader sequence. The expression and secretion of this gene in the yeast secretory pathway were expected. The first step of developing a yeast secretion vectors was to correct the reading frame of the multiple cloning sites on a general secretion vector pYA41. There are seven vectors derived from pYA41, viz: pYA41'/35, pYA41'/45, pUR 222/α1. pUC9/α1, pUC9/α2, pUC19/α1, pUC19/α2. These derivative secretion vectors have different reading frame on their multiple cloning sites which could be used in either sticky or blunt end ligation and can be manipulated to insert foreign genes. Secondly, these vectors were combined with several selective markers and replicative origins of yeast. In this step, seven secretion vectors were constructed, viz: YAUT0, YAUT1, YAUT2, pUC9/α1S, YAMTS, YAUTS, YAMKS; fusions between the cloned yeast α-factor promoter-leader sequence and preS2-SAg gene had been constructed on the last four vectors. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/75578 |
全文授權: | 未授權 |
顯示於系所單位: | 植物科學研究所 |
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