title: Production of live offspring with predicted genotypes using PCR-RFLP analysis of polar bodies from mouse oocytes.
abstract: Accurate identification of genotypes in gametes and early embryos could facilitate the efficient production of offspring with desirable traits. This study demonstrates the feasibility of producing offspring with predictable genotypes from micromanipulated mouse oocytes. The Polymerase Chain Reaction (PCR) was used to amplify genes in the IA subregion of the major histocompatibility complex of the mouse. The validity of the approach was demonstrated in experiment 1 with IA haplotypes of unfertilized mouse ova amplified via PCR and distinguished by restriction fragment length polymorphism (RFLP) analysis. In experiment 2, fertilized oocytes were micromanipulated to remove the first and second polar bodies, which were then genotyped by validated PCR-RFLP procedures. Primary oocytes of heterozygous females contain two copies of each of the different alleles. Following meiosis I and II, the genotype of the ovum was predicted by subtracting the alleles observed in micromanipulated polar body samples. Sixty-two fertilized ova were micromanipulated and transferred to recipient females resulting in 27 live offspring (44%). The correct maternal contribution to the embryonic genotype was predicted in 19 of 27 (71%) offspring as confirmed by PCR-RFLP analysis of DNA from pup tails. Predicted genotypes of two pups were not confirmed (7%), whereas no prediction could be made in six cases (22%).
authors.affiliation: Department of Animal Sciences, University of Illinois, Urbana 61801, USA.
authors.forename: M B
authors.initials: MB
authors.lastname: Wheeler
authors.forename: J A
authors.initials: JA
authors.lastname: Noble
authors.forename: V L
authors.initials: VL
authors.lastname: Jarrell
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D001483:Base Sequence; D017931:DNA Primers / Q000235:genetics; D004624:Embryo Transfer; D005260:Female; D005821:Genetic Techniques; D005838:Genotype; D000949:Histocompatibility Antigens Class II / Q000235:genetics; D008297:Male; D051379:Mice; D008809:Mice, Inbred C3H; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D009865:Oocytes / Q000378:metabolism* / Q000648:ultrastructure*; D010312:Parthenogenesis / Q000235:genetics; D016133:Polymerase Chain Reaction; D012150:Polymorphism, Restriction Fragment Length; D011247:Pregnancy
chemical_list: D017931:DNA Primers; D000949:Histocompatibility Antigens Class II