Cell Culture Process Operations for Recombinant Protein Production

2013 ◽  
pp. 35-68 ◽  
Author(s):  
Susan Abu-Absi ◽  
Sen Xu ◽  
Hugh Graham ◽  
Nimish Dalal ◽  
Marcus Boyer ◽  
...  
Keyword(s):  
Cell Culture ◽  
Recombinant Protein ◽  
Recombinant Protein Production ◽  
Protein Production ◽  
Cell Culture Process ◽  
Culture Process ◽  
Process Operations

scholarly journals Polar Fixation of Plasmids during Recombinant Protein Production in Bacillus megaterium Results in Population Heterogeneity

2015 ◽  
Vol 81 (17) ◽  
pp. 5976-5986 ◽  
Author(s):  
Karin M. Münch ◽  
Johannes Müller ◽  
Sarah Wienecke ◽  
Simone Bergmann ◽  
Steffi Heyber ◽  
...  
Keyword(s):  
Cell Culture ◽  
Cell Division ◽  
Recombinant Protein ◽  
Bacillus Megaterium ◽  
Recombinant Protein Production ◽  
Protein Production ◽  
Population Heterogeneity ◽  
Exponential Growth Phase ◽  
Scale Production ◽  
Content Type

ABSTRACTDuring the past 2 decades,Bacillus megateriumhas been systematically developed for the gram-per-liter scale production of recombinant proteins. The plasmid-based expression systems employed use a xylose-controlled promoter. Protein production analyses at the single-cell level using green fluorescent protein as a model product revealed cell culture heterogeneity characterized by a significant proportion of less productive bacteria. Due to the enormous size ofB. megaterium, such bistable behavior seen in subpopulations was readily analyzed by time lapse microscopy and flow cytometry. Cell culture heterogeneity was not caused simply by plasmid loss: instead, an asymmetric distribution of plasmids during cell division was detected during the exponential-growth phase. Multicopy plasmids are generally randomly distributed between daughter cells. However,in vivo andin vitroexperiments demonstrated that under conditions of strong protein production, plasmids are retained at one of the cell poles. Furthermore, it was found that cells with accumulated plasmids and high protein production ceased cell division. As a consequence, the overall protein production of the culture was achieved mainly by the subpopulation with a sufficient plasmid copy number. Based on our experimental data, we propose a model whereby the distribution of multicopy plasmids is controlled by polar fixation under protein production conditions. Thereby, cell lines with fluctuating plasmid abundance arise, which results in population heterogeneity. Our results provide initial insights into the mechanism of cellular heterogeneity during plasmid-based recombinant protein production in aBacillusspecies.

Large-Scale Insect Cell-Culture for Recombinant Protein Production

1988 ◽  
Vol 6 (12) ◽  
pp. 1406-1410 ◽  
Author(s):  
Brian Maiorella ◽  
Duane Inlow ◽  
Andrea Shauger ◽  
David Harano
Keyword(s):  
Cell Culture ◽  
Recombinant Protein ◽  
Insect Cell ◽  
Recombinant Protein Production ◽  
Large Scale ◽  
Protein Production ◽  
Scale Insect ◽  
Insect Cell Culture
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