The Drosophila Schneider S2 (S2) Expression System enables expression of recombinant proteins constitutively, as well as inductively. This system can establish both transient and stable transformants with various selection markers. The generation of stable cell lines for increased expression or large scale expression of the desired protein is currently accomplished by cotransfection of both the expression and selection vectors. The selection vectors, pCoHYGRO and pCoBLAST, are commercially available using hygromycin-B and blasticidin S, respectively. Recently, we generated a plasmid, pCoPURO, for selection of transfected S2 cells using puromycin, which allows significant acceleration of the selection time. Although co-transfection of the selection marker with the plasmid for heterologous protein expression is functional in stable expression at short culture periods, the expression levels of stable transformants are continuously decreased during long culture times. To overcome this limitation, we generated pMT-PURO, a new plasmid that contains both the expression cassette and puromycin selection marker in a single plasmid. This system allows rapid selection and maintenance of the transformed S2 lines for extended culture periods.

译文

果蝇Schneider S2 (S2) 表达系统可组成性和诱导性表达重组蛋白。该系统可以通过各种选择标记建立瞬态和稳定的转化子。目前,通过共转染表达和选择载体来实现用于所需蛋白质的增加表达或大规模表达的稳定细胞系的产生。选择载体pCoHYGRO和pCoBLAST分别使用潮霉素B和blasticidin S可商购。最近,我们生成了一个质粒pCoPURO,用于使用嘌呤霉素选择转染的S2细胞,这可以显着加快选择时间。尽管将选择标记物与用于异源蛋白表达的质粒共转染在短培养期间具有稳定表达的功能,但在长培养时间内,稳定转化体的表达水平会持续降低。为了克服这一限制,我们产生了pMT-PURO,这是一种新的质粒,在单个质粒中同时包含表达盒和嘌呤霉素选择标记。该系统可以快速选择和维护转换后的S2品系,以延长培养时间。

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