Practice question

Generation of knockout mice by ES cell method follows ordered protocol. Homology arms plus positive-negative selection markers are cloned into targeting vector containing floxed or null allele. Vector electroporated into cultured embryonic stem cells where homologous recombination occurs. G418 and ganciclovir selection enriches correctly targeted clones confirmed by Southern blot analysis. Targeted ES cells microinjected into blastocysts implanted into pseudopregnant foster mothers produce chimeric offspring. Chimeras bred for germline transmission yield heterozygous knockout founders, later intercrossed to homozygous knockout lines.