These microinjected embryos are then implanted into the oviduct of pseudopregnant mice. If skillfully performed, three weeks later a mouse is born with the selected gene. Cross-breeding subsequently produces a continuing lineage of mice bearing the human microinjected gene. Animals developing from microinjected cells contain the foreign gene in all somatic cells and also in germ-line cells if the foreign gene was integrated into the recipient gene before the first cell division. Animals developing after gene transfer are always hemizygous because the transgene is not integrated into both copies of a homologuous pair of chromosomes.
Homozygosity is achieved by crossing two hemizygous animals.
Micromanipulation to create a microinjected line Gene Targeting As noted, microinjected animals are produced by the addition of a gene to pronuclear embryos. ES cells used for gene targeting are derived from the inner cell mass of blastocysts that often were harvested from one of the many inbred 129 strains. Genomic DNA used in gene targeting should be derived from the same strain of mouse as the ES cells used. The chimeric blastocysts are then transferred surgically into the reproductive tracts of pseudopregnant recipient female mice and brought to term. Chimeric pups may be recognized visually, by observing coat color patterns, within about a week after birth.