Engineering of recombinant Chinese hamster ovary cells using proteomic investigation = 프로테오믹스 연구를 통한 재조합 CHO 세포의 개량

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In the culture of recombinant Chinese hamster ovary (rCHO) cells, optimization of culture environmental parameters, such as low temperature, hyperosmolarity, culture pH, and sodium butyrate addition has been widely studied. Although optimizing culture conditions generally takes less time and labor than genetic engineering, it also has some side-effects including decrease of cell growth, viability, or culture longevity. Accordingly, my investigations to improve cell performance were focused on an alternative method, genetic engineering. Selection of target proteins for genetic engineering was carried out using a proteomic approach which finds the changes in protein expression according to changes of culture conditions. Sodium butyrate (NaBu) is known to enhance the specific productivity of Chinese hamster ovary cells expressing human thrombopoietin. In order to better understand the intracellular responses of these cells resulting from NaBu treatment, the proteomic profiles of cells treated with various concentrations of NaBu (0-3mM) were compared using two-dimensional electrophoresis (2-DE). Based on spot intensities, 80 high intensity protein spots were selected. Fifty-six of the 80 protein spots, which represent 28 different kinds of proteins, were identified by MALDI-TOF-MS and MS/MS. Compared to control without NaBu treatment, the expression levels of 2 proteins (glucose regulated protein 78 (GRP 78) and peroxiredoxin 4) were increased over two fold with NaBu treatment and the expression level of phosphopyruvate hydratase was decreased over two fold with NaBu treatment. Due to multiplicity (multiple spots for one protein), a change in one single spot intensity from a 2-DE gel image may not represent the total change in expression level for that protein. Western blot analyses of GRP78, HSC70 and ERp57 confirmed the results of the MS analyses, though a degree of change in expression level differed between the two methods. Accordingly, the proteomic approa...
Advisors
Lee, Gyun-Minresearcher이균민researcher
Description
한국과학기술원 : 생명과학과,
Publisher
한국과학기술원
Issue Date
2009
Identifier
327726/325007  / 020037308
Language
eng
Description

학위논문(박사) - 한국과학기술원 : 생명과학과, 2009. 8., [ ix, 110 p. ]

Keywords

CHO cells; proteomics; genetic engineering; 2D gel system; DIGE; CHO 세포; 프로테오믹스; 유전자 공학 기술; 이차원 전기영동; DIGE; CHO cells; proteomics; genetic engineering; 2D gel system; DIGE; CHO 세포; 프로테오믹스; 유전자 공학 기술; 이차원 전기영동; DIGE

URI
http://hdl.handle.net/10203/27675
Link
http://library.kaist.ac.kr/search/detail/view.do?bibCtrlNo=327726&flag=dissertation
Appears in Collection
BS-Theses_Ph.D.(박사논문)
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