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postgraduate thesis: Optimization of conditions for expression and purification of recombinant human adiponectin
Title | Optimization of conditions for expression and purification of recombinant human adiponectin |
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Authors | |
Issue Date | 2016 |
Publisher | The University of Hong Kong (Pokfulam, Hong Kong) |
Citation | Chan, K. [陳筠梃]. (2016). Optimization of conditions for expression and purification of recombinant human adiponectin. (Thesis). University of Hong Kong, Pokfulam, Hong Kong SAR. |
Abstract | Obesity is one of the significant risk factors for the chronic diseases development, and chronic diseases are the leading cause of mortality globally. These chronic diseases are including Type 2 Diabetes Mellitus, atherosclerosis, and cardiovascular diseases. The studies of adipose tissues indicate its major role in energy homeostasis, glucose, and fat metabolism. Adiponectin, one of the adipokines synthesized from adipose tissues, shows the beneficial effects on promoting weight loss, improving insulin sensitivity, reducing the risk of atherosclerosis, and function of anti-inflammation.
In this study, we optimize the protein expression in Escherichia coli BL21 (DE3) cells by decreasing the induction temperature from 37°C to 25°C with extended induction time up to 18 hours to promote proper folding. Also, the concentration of inducer Isopropyl β-D-thiogalactoside was decreased from 0.5 mM to 0.2 mM to minimize the potential protein toxicity side effects. In addition, a step-gradient of imidazole elution buffer was used in hexahistidine-tagged protein purification by immobilized metal-ion affinity chromatography to promote a high purity level of the target protein. As a result, a high quantity and purity of recombinant human adiponectin were synthesized; that is 4.8 mg of recombinant human adiponectin from a 500 ml culture medium and with a high purity level of about 90% shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis. |
Degree | Master of Medical Sciences |
Subject | Adipose tissues |
Dept/Program | Medicine |
Persistent Identifier | http://hdl.handle.net/10722/237188 |
HKU Library Item ID | b5804688 |
DC Field | Value | Language |
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dc.contributor.author | Chan, Kwan-ting | - |
dc.contributor.author | 陳筠梃 | - |
dc.date.accessioned | 2016-12-28T02:01:44Z | - |
dc.date.available | 2016-12-28T02:01:44Z | - |
dc.date.issued | 2016 | - |
dc.identifier.citation | Chan, K. [陳筠梃]. (2016). Optimization of conditions for expression and purification of recombinant human adiponectin. (Thesis). University of Hong Kong, Pokfulam, Hong Kong SAR. | - |
dc.identifier.uri | http://hdl.handle.net/10722/237188 | - |
dc.description.abstract | Obesity is one of the significant risk factors for the chronic diseases development, and chronic diseases are the leading cause of mortality globally. These chronic diseases are including Type 2 Diabetes Mellitus, atherosclerosis, and cardiovascular diseases. The studies of adipose tissues indicate its major role in energy homeostasis, glucose, and fat metabolism. Adiponectin, one of the adipokines synthesized from adipose tissues, shows the beneficial effects on promoting weight loss, improving insulin sensitivity, reducing the risk of atherosclerosis, and function of anti-inflammation. In this study, we optimize the protein expression in Escherichia coli BL21 (DE3) cells by decreasing the induction temperature from 37°C to 25°C with extended induction time up to 18 hours to promote proper folding. Also, the concentration of inducer Isopropyl β-D-thiogalactoside was decreased from 0.5 mM to 0.2 mM to minimize the potential protein toxicity side effects. In addition, a step-gradient of imidazole elution buffer was used in hexahistidine-tagged protein purification by immobilized metal-ion affinity chromatography to promote a high purity level of the target protein. As a result, a high quantity and purity of recombinant human adiponectin were synthesized; that is 4.8 mg of recombinant human adiponectin from a 500 ml culture medium and with a high purity level of about 90% shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis. | - |
dc.language | eng | - |
dc.publisher | The University of Hong Kong (Pokfulam, Hong Kong) | - |
dc.relation.ispartof | HKU Theses Online (HKUTO) | - |
dc.rights | This work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License. | - |
dc.rights | The author retains all proprietary rights, (such as patent rights) and the right to use in future works. | - |
dc.subject.lcsh | Adipose tissues | - |
dc.title | Optimization of conditions for expression and purification of recombinant human adiponectin | - |
dc.type | PG_Thesis | - |
dc.identifier.hkul | b5804688 | - |
dc.description.thesisname | Master of Medical Sciences | - |
dc.description.thesislevel | Master | - |
dc.description.thesisdiscipline | Medicine | - |
dc.description.nature | published_or_final_version | - |
dc.identifier.doi | 10.5353/th_b5804688 | - |
dc.identifier.mmsid | 991020889659703414 | - |