Stability genetic

As it is imperative that the plant-derived hiopharmaceutical product must be obtained repeatedly and on a consistent basis, a master cell culture bank, seed bank for transgenic plants, or virus seed stock for transient expression systems must be constantly maintained. Storage conditions must therefore he optimized to prevent contamination and ensure viability. Both transgene stability (e.g., reversion to wild type or sequence drift of plant virus expression vectors) and protein expression levels must be monitored in a representative plant of a given bank or stock to minimize any possible variation in expression levels that may affect safety and consistency of the hnal product. A program that monitors lot-to-lot consistency of the hiochemical and biological properties by comparing the product with appropriate in-house reference standards could he implemented as a fundamental component of product development. 

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Pillai SD, Pepper IL (1991) Transposon Tn5 as an identifiable marker in rhizobia survival and genetic stability of Tn5 mutant bean rhizobia under temperature stressed conditions in desert soils. Microbial Ecol 21 21-33 Pote J, Ceccherini MT, Van VT, Rosselli W, Wildi W, Simonet P, Vogel TM (2003) Fate and transport of antibiotic resistance genes in saturated soil columns. Eur J Soil Biol 39 65-71... 

There are many uncertainties in the use of bioassay methods, not the least of these being the genetic stability of the assay organisms. Stock cultures cannot be treated like chemical reagents, to be put back on the shelf and forgotten between analytical runs. This is an area of analysis best left to the microbiologists if the information is absolutely necessary, the maintenance of cultures and the actual assay should not be left solely in the hands analytical chemists. 

Besman M.J. and Shiba D. (1997), Evaluation of genetic stability of recombinant Human Factor VIII by peptide mapping and on-line mass spectrometric analysis. Pharm. Res. 14(8), 1092-1098. 

In the North Sea project, the results from primary screening for biological activity or new compounds guide the selection of strains for upscaling and finally isolation and structural elucidation. Since even modern methods for structure determination and an initial biomedical evaluation require 10 mg of every compound or more, a scale-up fermentation is necessary. With the aid of biotechni-cal methods, fermentation conditions have to be optimized to achieve maximum yield of metabolites, to increase the genetic stability of the producer or to improve other parameters. 

Mutagenic potential—effects on genetic stability of bacteria (Ames test) of mammalian cells in culture... 

Figure 4.9. Optimization of product yieid through sequential and systematic screening and selection of genetic construct and recombinant host cells that produce the highest yield and genetic stability. The schematically presented steps are required to develop a master cell-bank, and working stock for the production of biologically active recombinant proteins in pharmaceutical scale.

One consideration to bear in mind during the design of inoculums expansion is to demonstrate the genetic stability of the cell line beyond the expected number of generations required to operate at large-scale. This is usually accomplished by conducting measurements of product expression and genetic markers in cells from an extended cell bank (ECB). 

Mutagenic potential Test effects on genetic stability and mutations in bacteria (Ames test) or mammalian cells in culture dominant lethal test and clastogenicity in mice. 

MSCs offer several advantages for the treatment of GVHD and inflammatory and immune disorders. The ease of isolation, in vitro expansion, genetic stability, and ability to escape alloimmunity, make MSCs a model of choice for cellular therapy and also for gene... 

One method used to achieve genetic stability is to insert the plasmid or the recombinant DNA directly into the chromosome. Since no cell can afford to lose a chromosome, this assures that the recombinant DNA is not lost as long as it remains an integral part of the chromosome. 

Radman, M., Wagner, B.W., Glickman, B.W., Meselson, M. (1980). DNA methylation, mismatch correction and genetic stability. In Progress in Environmental Mutagenesis (Alacevic, M., Ed.), pp. 121-130. Elsevier/Nth. Holland Biomedical, Amsterdam. 

Peptide mapping is a method that enables the determination of protein identity when compared to a standard. When compared to previous lots of the same product, it serves to determine the stability of the protein s primary sequence, which in turn reflects the genetic stability of the producer cells. This method is capable of detecting small differences between proteins in one or more amino acids. The detection will be dependent upon an amino acid alteration affecting the observed peptide profile (Figure 13.1 see color section). 

Ensuring long term genetic stability of cells... 

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