Random integration

Davies, J. and Reff, M., Chromosome localization and gene-copy-number quantification of three random integrations in Chinese-hamster ovary cells and their amplified cell lines using fluorescence in situ hybridization, Biotechnol. Appl. Biochem., 33, 99-105, 2001. 

Classical or Conventional Transgenic Mice (Tg) Random Integration of the Transgene in the Mouse Genome... 

Fig. 1. Outline of the generation of transgenic mice via pronuclear microinjection. Transgenic animals carry the transgenes randomly integrated (hemizygous, tandems, dominant).

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 concern over integration of pDNA into the host chromosomal pDNA is a significant safety concern since random integration could result in the activation of a proto-oncogene. To date, in preclinical studies, there is no direct evidence that pDNA integrates into chromosomal DNA following i.m. administration (Nichols et al., 1995 Martin et al., 1999 Ledwith et al., 2000 Manam et al., 2000). It has been determined that if such an event were to occur, it would be well below the level of the natural spontaneous mutation rate and would not pose a significant safety concern (Martin et al., 1999). 

The predominance of episomal persistence in latent rAAV infection has a number of important implications. First, it helps to explain why rAAV gene transfer has been particularly effective in terminally differentiated nonproliferating cell populations. In these cells episomal forms are quite stable and are not diluted out by host cell division. A second important aspect of episomal persistence is that this mode of persistence decreases the risk of insertional mutagenesis that might otherwise be associated with random integration of vector DNA into the host cell genome. 

Retrovirus 7 kb Long-term expression Random integration into host chromosome Early Can only transduce dividing cells... 

Lenti virus 10 kb Long-term expression Random Integration into host chromosome Early Insertional mutagenesis concerns... 

Retrovirus has RNA genome integrate into host chromosomes sustained gene expression incorporate up to 8kb of ON A do not infect post-mitotic cells random integration problematic large-scale production possible insertional mutagenesis... 

Fig. 7.1. The figure illustrates the hypothetical results obtained from three different cell clones after random integration of a marker gene, following either plasmid transfection or retroviral infection. Genomic DNA digestion with the restriction enzymes A, B or C will result in a unique band pattern, which is suggestive of the integration events in each cell clone.

The integration of HIV into the host chromosome is troublesome for several reasons. First, HIV can establish a chronic and persistent infection, particularly in long-lived cells of the immune system such as memory T-lymphocytes. ° Second, integration is random, thus making it difficult to target and extract integrated HIV. Last, random integration of HIV may cause cellular abnormalities and induce apoptosis. 

---