کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
7234845 1471012 2018 10 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Single-cell cloning enables the selection of more productive Drosophila melanogaster S2 cells for recombinant protein expression
موضوعات مرتبط
مهندسی و علوم پایه مهندسی انرژی انرژی های تجدید پذیر، توسعه پایدار و محیط زیست
پیش نمایش صفحه اول مقاله
Single-cell cloning enables the selection of more productive Drosophila melanogaster S2 cells for recombinant protein expression
چکیده انگلیسی
The generation of monoclonal cell lines is an important early process development step for recombinant protein production. Although single-cell cloning is an established method in mammalian cell lines, straightforward protocols are not yet available for insect cells. We describe a new method for the generation of monoclonal insect cells without using fetal bovine serum and/or feeder cells pretreated by irradiation or exposure to mitomycin. Highly productive clones of Drosophila melanogaster S2 cells were prepared in a two-step procedure, comprising the establishment of a polyclonal population and subsequent single cell isolation by limiting dilution. Necessary growth factors were provided by co-cultivation of single transformants with untransfected feeder cells, which were later removed by antibiotic selection. Enhanced expression of EGFP and two target peptides was confirmed by flow cytometry and dot/western blotting. Highly productive clones were stable, showed a uniform expression profile and typically a sixfold to tenfold increase in cell-specific productivity.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Biotechnology Reports - Volume 19, September 2018, e00272
نویسندگان
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