title: Rapid detection of male-specific DNA sequence in bovine embryos using fluorescence in situ hybridization.
abstract: An accurate, reliable, and quick (less than an hour) method for determining the sex of bovine embryos was developed using a fluorescence in situ hybridization (FISH), with a probe designed from a bovine Y chromosome specific DNA (BC1.2). First, to improve a protocol of FISH and evaluate an accuracy of the method, lymphocyte nuclei prepared from three bulls, two cows, and one freemartin were tested. We found that 5 min was enough for hybridization. The washing solution adequate for posthybridization was 0.5x SSC at 72 degrees C for 5 min. The whole procedure for FISH can be accomplished in less than an hour. A male-specific signal was detected, on average, as 97, 0.5, and 83%, respectively, of lymphocytes in males, females, and a freemartin. Using the rapid FISH protocol developed, 28 embryos were divided. According to the presence of the digoxigenin signal, 16 embryos (57.1%) were predicted as male, and 12 embryos (42.9%), predicted as female.
authors.affiliation: Miyagi Agricultural College, Sendai, Japan.
authors.forename: J
authors.initials: J
authors.lastname: Kobayashi
authors.forename: A
authors.initials: A
authors.lastname: Sekimoto
authors.forename: H
authors.initials: H
authors.lastname: Uchida
authors.forename: T
authors.initials: T
authors.lastname: Wada
authors.forename: K
authors.initials: K
authors.lastname: Sasaki
authors.forename: H
authors.initials: H
authors.lastname: Sasada
authors.forename: M
authors.initials: M
authors.lastname: Umezu
authors.forename: E
authors.initials: E
authors.lastname: Sato
mesh_terms: D000818:Animals; D002417:Cattle; D002467:Cell Nucleus / Q000737:chemistry; D004247:DNA / Q000737:chemistry*; D004622:Embryo, Mammalian / Q000737:chemistry*; D005260:Female; D005611:Freemartinism / Q000235:genetics; D017404:In Situ Hybridization, Fluorescence / Q000379:methods / Q000662:veterinary*; D008214:Lymphocytes / Q000737:chemistry; D008297:Male; D017422:Sequence Analysis, DNA / Q000379:methods / Q000662:veterinary; D012727:Sex Characteristics; D012732:Sex Determination Analysis / Q000662:veterinary*; D019849:Sex Determination Processes; D014998:Y Chromosome
chemical_list: D004247:DNA