Production of tpa using eukaryotic n prokaryotic cells Essay - 1

Production of tpa using eukaryotic n prokaryotic cells - Essay Example course of production, safety precautions have to be strictly adhered to, as the drugs are poisons in nature and could cause severe body harm if misused. A method for producing tissue plasminogen activator (t-PA) in eukaryotic host cells is disclosed. Enhanced levels of t-PA production are obtained by co-amplification of the t-PA gene through treatment of cultures transformed with mutant or wild type DHFR with methotrexate. Â  A cell culture comprising methotrexate (MTX) sensitive recombinant host cells transformed with an expression vector comprising a first DNA sequence encoding a dihydrofolate reductase (DHFR) protein with a low binding affinity for MTX, and a second DNA sequence encoding human tissue plasminogen activator (tPA), tPA encoded by said second DNA sequence, and an effective amplifying concentration of MTX. The invention herein relates to the production of human tissue plasminogen activator (tPA) in a transformant host cell culture. More specifically, the invention relates to vectors, cells, and methods of producing tPA in conjunction with expression of the sequences for coding for dihydrofolate reductase (DHFR) protein in such cells. Another example that describes use of CHO cells as host cells, and expression vectors which include the SV40 origin of replication as a promoter. However, it would be well within the skill of the art to use analogous techniques to construct expression vectors for expression of desired protein sequences in alternative eukaryotic host cell cultures. If cells without formidable cell wall barriers are used as host cells, transfection is carried out by the calcium phosphate precipitation method as described by Graham and Van der Eb, Virology, 52:546 (1978). However, other methods for introducing DNA into cells such as by nuclear injection or by protoplast fusion may also be used. Construction of suitable vectors containing the desired coding and control sequences employ standard ligation techniques. Isolated plasmids or