Sara Yousefi Taemeh; Jalil Mehrzad; Hesam Dehghani
Abstract
Primordial germ cells (PGCs) are precursors of mature gametes, which transmit genetic information to the next generation. Due to the importance of PGCs in many fields, including developmental biology, genome editing, transgenesis, and conservation of avian genetic resources, various research aspects ...
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Primordial germ cells (PGCs) are precursors of mature gametes, which transmit genetic information to the next generation. Due to the importance of PGCs in many fields, including developmental biology, genome editing, transgenesis, and conservation of avian genetic resources, various research aspects have focused on the cultivation of PGCs. Despite considerable progress in the establishment of specified culture media for the expansion of PGCs, a well-defined PGC culture medium has not yet been developed. This might be due to the complexity of the nutritional requirements of PGCs in the culture. Besides the nutritional needs, including vitamins, amino acids, salts, carbohydrates, and growth factors, a particular source of energy must be provided to sustain growth and viability. Glutamine is a major energy source for cultured cells, commonly added in cell culture media at higher concentrations than other amino acids. However, glutamine is very labile and rapidly degrades in solutions such as culture media. This generates ammonia as a by-product, which is toxic to the cultured cells and can affect cell viability and protein glycosylation. Therefore, the stability of glutamine in culture conditions is another concern for the long-term culture of PGCs. Here, we study the effect of glutamine stability on PGC culture using glutamine and GlutaMax (a commercial stabilized dipeptide form of glutamine). We found that the addition of GlutaMax in the medium promotes PGC proliferation. This effect might be exerted by minimizing production of toxic ammonia that results in maximizing cell performance and media stability.
Mohsen Naeemipour; Mohammadreza Bassami
Abstract
Nowadays, production of recombinant proteins in eukaryotes is gaining good deal of attention. Transgenic chicken as a eukaryotic system has a high potential for producing recombinant proteins. Post-translational changes, especially glycosylation, are characteristic of the eukaryotic proteins. In practice ...
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Nowadays, production of recombinant proteins in eukaryotes is gaining good deal of attention. Transgenic chicken as a eukaryotic system has a high potential for producing recombinant proteins. Post-translational changes, especially glycosylation, are characteristic of the eukaryotic proteins. In practice we need to choose a proper expressing host when considering over-expression of a recombinant protein. Chickens are among the well-considered candidates for such application. Production of transgenic chickens could be achieved in different ways, including application of primordial germ cells. Primordial germ cells are progenitor of sperm and ovum. These cells are round, with a big nucleus and a cytoplasm with lipid and glycogen particles. The first step for having transgenic chickens is isolation and culture of the primordial gem cells. In the present study, these cells were isolated by centrifugation method in presence of ficoll and using magnetic cell sorting, and were cultured in optimal culture medium. These cells were finally characterized with defined methods, like Periodic acid-schiff staining, alkaline phosphates activity assessment, and antibody staining.
