中 文 摘 要

本基因重組菌為JM109大腸桿菌內殖入載體pQE-51-pSCR所構成，而此載體內含Pseudomonas putida NTU-8肌酸酶基因及前端插入一段來自於Aeromonas hydrophila幾丁質分解酵素的信號序列，此基因轉殖菌因質體具有幾丁質酵素的信號序列，所以可將肌酸酵素分泌至細胞間質。

本論文以LB培養基與將其中部分tryptone與yeast extract以3克、6克和9克葡萄糖置換的複合培養基進行醱酵培養，目的在於探討IPTG誘導時間對於個別培養基中菌體濃度與肌酸酵素活性變化之影響。實驗中發現不論在LB或其餘3個添加葡萄糖的複合培養基，在適當的時間誘導皆可促進菌體生長，推測原因可能為誘導後加速某特定胺基酸之利用效率所造成。為確定此構想，而嘗試分析與TCA cycle較接近的絲胺酸、丙胺酸、天門冬胺酸與麩胺酸在醱酵液中的濃度，以進一步瞭解誘導前後其濃度的變化。

ABSTRACT

The recombinant Escherichi coli was constructed by inserting the pQE-51-pSCR01 expression vector in Escherichia coli JM109. It can express the chitinase signal peptide – creatinase hybrid gene by way of the expression vector. Besides, the target protein can be excreted to the periplasmic space due to the fusing of the signal peptide.

We processed the fermentations by incubating the recombinant Escherichi coli in Luria-Bertani medium and the media with parts of tryptone and yeast extract replaced by glucose. Different behaviors for the cell growth and creatinase productivity in various media were found if the inducer of isopropyl-β-D-thiogalactopyranoside (IPTG) was introduced at different periods of time. The cell concentration and creatinase activity were enhanced when inducing IPTG at an appropriate time. Possible explanation for such enhancements was attributed to increasing usage of certain amino acid. For confirming this hypothesis, the analysis of the concentration of serine、aspartic acid、alanine and glutamic acid close to TCA cycle was performed.

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