Gene transfer into mouse embryos

Abstract

Gene transfer into the murine genome was accomplished nearly a decade ago by use of chimeras and teratocarcinomas; however, the low frequencies of transfer into the germ line and other difficulties stemming from mosaicism and karyotypic abnormalities in chimeric mice have limited the general usefulness of this procedure in achieving transformation in mammalian embryos. The introduction of cloned genes into teratocarcinoma cells, selection for a mutant phenotype, and transfer of those cells into mouse embryos holds some promise as a technique to employ mouse chimeras for gene transfer into mice. Infection with animal viruses and retroviral vectors provides another way to introduce exogenous DNA into mouse embryos. Infection with Mo-MuLV has been utilized to characterize the relationship between sites of integration and gene function in developing and adult mice. Gene transfer by microinjection of cloned recombinant DNA has been used by many laboratories for the transfer of DNAs into mouse embryos. The factors affecting transformation frequencies and sites of integration are unknown at present, although it seems that integration is not strictly mediated by homology-dependent events. Many genes have been introduced into mouse embryos by these procedures and many of these are expressed at high levels in appropriate tissues. No realistic possibility exists at the present time for the utilization of embryo gene transfer in the medical field for the correction of genetic defects for several reasons. First, in order to effectively provide "gene therapy" it would be necessary to determine the genotype of each recipient egg, a technical impossibility. The genetic diseases that would be amenable to germ line intervention are recessive diseases and there would be only a 25% chance of any one embryo derived from heterozygous parents being a homozygous recessive. Moreover, it would be impossible to distinguish the normal from abnormal embryos. Second, the frequencies of transformation are so low as to exclude work on human beings on ethical grounds. Third, the parameters effecting chromosomal integration sites and gene expression have not been fully characterized. Until it becomes experimentally possible to target the newly introduced DNA into expressable chromosomal sites and actively replace or supplement defective genes, the possibility of gene therapy through manipulation of embryos is remote. Yet, efforts to provide gene therapy in somatic tissues have been promising, leading to expression of a modified phenotype (Anderson, 1984). In contrast to embryo gene therapy, gene therapy in somatic tissues would not lead to germ line propagation of the manipulated genotype.(ABSTRACT TRUNCATED AT 400 WORDS)