Mammalian cells expression purification

The third time constraint depends on whether the product can be extracted from seeds or fruits. This uncouples protein expression and purification. Large batches of seeds containing the recombinant protein can be produced and stored at low costs. Provided the protein remains stable in the stored seeds, purification can be carried out on demand or shifted according to free capacities. The advantage of one large harvest, with seeds mixed to uniformity, is that this allows production on demand. In contrast, mammalian cell culture is prone to minor batch-to-batch variations in... 

Makrides SC (1999), Components of vectors for gene transfer and expression in mammalian cells, Protein Expr. Purif. 17 183-202. 

Rasmussen LK, Larsen YB, Hojrup P (2005), Characterization of different cell culture media for expression of recombinant antibodies in mammalian cells presence of contaminating bovine antibodies, Protein Expres. Purif. 41 373-377. 

The rDNA-derived products may contain potentially harmful contaminants that are normally not present in their equivalents prepared by chemical methods, and which the purification process must be capable of eliminating, such as the endotoxins expressed in bacterial cells, cellular DNA, and viruses of animal origin. Contamination with nucleic acid from transformed mammalian cells is of particular concern due to the possible presence of potentially oncogenic DNA. 

Generating suitable protein is often the most labor intensive step on the way to 3D structures. Enough protein to create hundreds of crystals is needed and therefore care needs to be taken with expression and purification procedures. High yield expression and simple and effective purification protocols are beneficial. Optimized protein constructs for crystallization often lack glycosylation sites and carry affinity tags for purification. All standard structural biology protein expression systems are used to produce the proteins for fragment cocrystallization, i.e., E. coli, Baculo virus insect cell systems or mammalian cell lines. 

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