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タイトル: New strategy for rapid isolation of stable cell lines from DNA-transformed insect cells using fluorescence activated cell-sorting
著者: Kato, Tatsuya
Yoshizuka, Kengo
Park, Enoch Y.
掲載誌名: Journal of Biotechnology
出版者: Elsevier
巻: 147
号: 2
開始ページ: 102
終了ページ: 107
出版日付: 2010-05-17
権利: Copyright © 2010 Elsevier B.V. All rights reserved.
NDC: 463
抄録: A stably transformed insect cell expression system is superior to a baculovirus expression system, since the expression system is sustained and there is no cell lysis, but the isolation of cell lines producing recombinant proteins is time-consuming and laborious. In this study, we developed a technique for the rapid isolation and efficient cultivation of sorted cells in a 24 well deep plate Bioshaker, utilizing the fluorescence activated cell-sorting (FACS) method. TnpXme11 cells, which stably expressed GFP(uv)-beta1,3-N-acetylglucosaminyltransferase2 (GGT2), were transfected with a plasmid vector for the expression of a molecular chaperone (TnpXme11-hCNX6 cell line). The expression levels of GGT2 and the molecular chaperone fused with HcRed were analyzed by FACS. Two cell lines were established by single and double sorting. Sorting of the top 10% of the TnpXme11-hCNX6 cell population with the highest fluorescence yielded the TnpXme11-hCNX6-1 cell line. TnpXme11-hCNX6-2 cells were created in a similar fashion, as mentioned above, by a second sorting of the top 10% of the TnpXme11-hCNX6-1 cell population with the highest fluorescence. The cells thus isolated produced approximately 2-fold higher extracellular activity than that before cell sorting. This procedure can be accomplished in only 2 weeks, including transfection, isolation and analysis of high protein-producing cells, and is a breakthrough strategy for the rapid isolation of a recombinant, stable insect cell line.
ISSN: 01681656 OPAC
E-ISSN: 18734863 OPAC
出版者DOI: 10.1016/j.jbiotec.2010.03.011   
PMID: 20347890
NII書誌ID: AA10458361 OPAC ciniib
バージョン: author
出現コレクション:11. 雑誌論文・記事(Journal Article, Article, Preprint)

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Others By: Kato, Tatsuya (加藤, 竜也) (カトウ, タツヤ) -- Yoshizuka, Kengo -- Park, Enoch Y. (朴, 龍洙) (パク, ヨンス)



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