碩士 / 國立中正大學 / 化學工程研究所 / 104 / Whole blood transfusions are limited by cross matching, disease transmission and blood storage. To develop a safe and effective red blood cell substitute is being a great challenge. Using Escherichia coli (E. coli)as a host to produce soluble recombinant human hemoglobin alpha and beta chains is attractive. Insufficient heme would let the structure of hemoglobin become unstable and easy to be degraded. However, the endogenous heme production from the non-pathogenic stain of E. coli is insufficient to supply the need for recombinant hemoglobin production. Additionally, the non-pathogenic stain of E. coli lacks of heme transporter that can transport extracellular heme inti cells for utilization. In order to increase the supply of heme and stabilize hemoglobin, we cloned chuA gene from the chromosome of E. coli O157: H7 into plasmid pET-29c and then subcloned the chuA gnen with the T7 promoter and terminator to plasmid pET-Hb(β67W-β95C-synoα-α29F)-MAP becoming pET-Hb(β67W-β95C -synoα-α29F)-MAP-chuA. When transformed into E. coli BL21(DE3) and JM109(DE3), plasmid pET-Hb(β67W-β95C-synoα-α29F)-MAP-chuA was able to express chuA and increased the cell growth. However, the introduction of chuA did not improve the hemoglobin expression. Moreover, the hemoglobin expression of E. coli JM109(DE3) is better than that in E. coli BL21(DE3).
| Identifer | oai:union.ndltd.org:TW/104CCU00063024 |
| Date | January 2016 |
| Creators | CHEN,YI-WEN, 陳怡文 |
| Contributors | HUANG,KUANG-TSE, 黃光策 |
| Source Sets | National Digital Library of Theses and Dissertations in Taiwan |
| Language | zh-TW |
| Detected Language | English |
| Type | 學位論文 ; thesis |
| Format | 82 |
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