Expression vectors integration

Several approaches have been developed to reduce the time needed to obtain high-yielding cell lines. These approaches exploit the importance of the chromosomal locus in determining the level of gene expression to increase the proportion of transfectants with the expression vector integrated into a transcriptionally active locus by up to 10-fold. Since these approaches generate a clone with only one or at most a few copies of the expression vector [1, 25], the problems associated with amplification are eliminated. 

Lundstrom, K. (2003) Semliki Forest vims vectors for rapid and high-level expression of integral membrane proteins. Biochimica et Biophysica Acta, 1610 (1), 90-96. 

Retrovirus RNA Transduction efficiency is high. Sustained expression of vector after integrating into host genome. Host does not express vector proteins Dividing cells are required for infectivity. Random integration. Oncogenesis may be induced after integration... 

The important contribution of these episomal forms to transgene expression has recently been demonstrated in mouse fiver (Nakai et al., 2001). In this study a partial hepatectomy was performed on mice after portal vein injection of a rAAV vector, in order to stimulate proliferation of the transduced fiver. In this situation one would predict that expression from integrated vector genomes would not decrease significantly, since the genome copy... 

Useful to confirm the integrity of expression vectors, convenient for high-throughput screening methods, e.g. to study different mutant genes Circular DNA is used... 

The key function of the vector is to introduce the cDNA under the control of a strong promoter and, if a stable cell line is to be developed, also to introduce resistance to a compound that is otherwise toxic to the host cell. This facilitates selection (only vector-bearing cells grown in special media) of a minority of cells that have incorporated the vector. There are many adequate expression vectors available from a number of commercial suppliers. Several classes of vector systems appear appropriate for transporter expression as integrating, episomal, retroviral, Vaccinia virus-, and Adenovirus-based systems have all been used successfully in studies referenced in this chapter. 

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