Microsurgically produced homozygous-diploid uniparental mice

Abstract

Shortly after fertilization, either the male or the female pronucleus was microsurgically removed from 202 F(1) hybrid eggs derived from crosses of two inbred strains. Subsequent incubation of these haploid eggs in medium containing cytochalasin B, which inhibits cytokinesis but not nuclear division, enabled the remaining pronucleus to become diploid. After nuclear diploidization and transfer to regular culture medium, cleavage commenced normally, and a total of 135 successfully manipulated eggs continued in development and yielded 93 morulae and blastocysts. These embryos were surgically transferred to the uteri of pseudopregnant foster mothers who gave birth to seven live female offspring. Five of the females were derived from the maternal genome (gynogenesis) and the remaining two mice inherited only the paternal genes (androgenesis), depending on whether the female or male pronucleus had been retained in the egg, respectively. Homozygosity for a number of genetic loci positioned on different chromosomes and effecting the coat color phenotype and strain-specific allelic variants of several enzymes, urinary and plasma proteins, and hemoglobins could be demonstrated unequivocally in all instances. Chromosomal analysis revealed a normal diploid karyotype including two X chromosomes. Thus far, six of the seven homozygous-diploid (isogeneic) females have proved to be fertile and have given birth to progeny corresponding only to the pronuclear genotype of the mother.