Handbook of Industrial Cell Culture: Mammalian, Microbial, by Victor A. Vinci, Sarad R. Parekh

By Victor A. Vinci, Sarad R. Parekh

Members from pharmaceutical, agriculture, biotechnology, and different businesses clarify equipment for culturing cells on a wide scale to generate advertisement items. Their issues contain genetic techniques to generating recombinant proteins in mammalian cells, method improvement issues for mammalian mobilephone tradition, genetic engineering options for traditional items in actinomycetes, tradition medium optimization and scale-up for microbial fermentations, oleosin partitioning know-how for generating recombinant proteins in oil seeds, proteomics, and DNA shuffling for whole-cell engineering.

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Interestingly, in their native environment highly specialized cells represent the most efficient mammalian expression systems known in the body, such as ß-globin synthesis in erythroblasts or immunoglobulin secretion by B-cells. Understanding the principles of gene expression in such cells could lead to recombinant cell lines with superior production properties. 1. Vector Design Generally, mammalian expression vectors are circular shuttle plasmid vectors (Fig. 1). Such plasmids usually carry a mammalian gene-expression cassette and an optional mammalian selection-marker gene.

37. Waldburger, J. , et al. (2000) Lessons from the bare lymphocyte syndrome: molecular mechanisms regulating MHC class II expression. Immunol. Rev. 178, 148–165. 38. Kaufman, R. , Wasley, L. , Spiliotes, A. , Gossels, S. , Latt, S. , Larsen, G. , et al. (1985) Coamplification and coexpression of human tissue-type plasminogen activator and murine dihydrofolate reductase sequences in Chinese hamster ovary cells. Mol. Cell Biol. 5, 1750–1759. 39. , et al. (1999) Engineering chinese hamster ovary cells to maximize sialic acid content of recombinant glycoproteins.

Similarly, the glycosylation pattern in BHK cells has been successfully rendered more human-like by expression of ␣2,6-sialyltransferase and 1,3-fucosyltransferase-III genes. Our understanding of the potential of glycosylation is incomplete. Although some general concepts have been clarified, important questions remain concerning activity and other relevant in vivo characteristics of individual glycoforms of a particular protein. Secreted protein products are a mixture of a number of different yet related isoforms.

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