Genotypic features of lentiviral transgenic mice

School of Life Sciences and "Frontiers in Genetics" National Center for Competence in Research, Ecole Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland; Ludwig Institute for Cancer Research, and Swiss Institute of Bioinformatics, 1015 Lausanne, Switzerland; Department of zoology and animal biology, University of Geneva, Geneva, Switzerland

^* To whom correspondence should be addressed. Email: didier.trono{at}epfl.ch.

	Abstract

Lentivector-mediated transgenesis is increasingly used, whether for basic studies as an alternative to pronuclear injection of naked DNA or to test candidate gene therapy vectors. In an effort to characterize the genetic features of this approach, we first measured the frequency of germ line transmission of individual proviruses established by infection of fertilized mouse oocytes. Seventy integrants from 11 founder mice were passed to 111 G1 pups for a total of 255 events, corresponding to an average rate of transmission of 44%. This implies that integration had most often occurred at the one- or two-cells stage, and that the degree of genotypic mosaicism in G0 mice obtained through this approach is generally minimal. Transmission analysis of 8 individual proviruses in 13 G2 mice obtained by G0-G1 cross revealed only 8% of proviral homozygocity, significantly below the 25% expected from purely mendelian transmission, suggesting counter-selection due to interference with the functions of targeted loci. Mapping of 239 proviral integration sites in 49 founder animals revealed that about sixty percent resided within annotated genes, with a marked tendency for clustering in the middle of the transcribed region, without influence of orientation. Transcript levels of a set of arbitrarily chosen target genes were significantly higher in 2-cell embryos than in embryonic stem cells or adult somatic cells, suggesting that, as previously noted in other settings, lentiviral vectors integrate preferentially into regions of the genome that are transcriptionally active or poised for activation.