Genetic manipulation by means of microcell-mediated transfer of normal human chromosomes into recipient mouse cells

Abstract

Microcell-mediated chromosome transfer is an innovative approach to the production of karyotypically simple hybrids. This method of gene transfer, employing micronuclei formed by prolonged Colcemid treatment, has been utilized for rodent systems. Expansion of this technology to include transfer of normal human genetic material has been hindered because large micronucleate populations from diploid human cells have been unobtainable. This report describes the production of micronuclei in 40-60% of normal human fibroblasts. These micronucleated cells have been enucleated by combining centrifugation and cytochalasin B treatment, and the resultant microcells have been purified and fused to recipient mouse (LMTK-) cells. Microcell hybrid clones containing a single human chromosome have been isolated from three separate fusion experiments. The time course for production of these hybrids, from fusion to karyotypic analysis, was 6 weeks. With a transfer frequency of about 2 x 10(-6), a single intact human chromosome has become a functioning element of the murine genome.