Process, vectors and engineered cell lines for enhanced large-scale transfection

摘要

Processes, vectors and engineered cell lines for large-scale transfection and protein production in mammalian cells, especially Chinese Hamster Ovary (CHO) cells are described in which transfection efficiencies are realized through the use of a single vector system, the use of functional oriP sequences in all plasmids, the use of codon-optimized Epstein-Barr virus nuclear antigen-1 (EBNA1) constructs, the use of a fusion protein between a truncated Epstein-Barr virus nuclear antigen-1c (EBNA1c) protein and a herpes simplex virus protein VP16, the use of a 40 kDa fully deacetylated poly(ethylenimine) as a transfection reagent, the use of co-expression of a fibroblast growth factor (FGF) and/or the use of protein kinase B to potentiate heterologous gene expression enhancement by valproic acid (VPA).

主权项

1. An expression system comprising one vector having an Epstein-Barr virus (EBV) nuclear antigen-1 (EBNA1) nucleotide sequence, a promoter and a polyadenylation signal for the EBNA1 nucleotide sequence, an EBV oriP nucleotide sequence, a gene of interest and a promoter and a polyadenylation signal for the gene of interest, wherein the expression system is integrated into a host genome and the gene of interest is stably expressed in a Chinese Hamster Ovary (CHO) cell.