Sexing of porcine embryo by in situ hybridization using chromosome Y- and 1-specific DNA probes

This study was carried out to determine if a rapid, simultaneous detection system using chromosome Y- and 1-bearing boar spermatozoa was applicable fo r sexing embryos. Porcine embryos were recovered from gilts and sows 4 to 6 d after mating, and whole embryos or biopsy cells were mounted on a glass slide with a small amount of fixative (methanol: acetic acid: distilled wat er = 9:1:4). The samples were then stained by means of a fluorescence in si tu hybridization (FISH) procedure developed specifically for the detection of Y-bearing spermatozoa. Hybridization was performed using digoxigenin (di g)-labeled chromosome Y- specific DNA, and biotin-labeled chromosome 1-spec ific DNA sequences were detected as a signal of FITC and Texas Red on nucle us visualized DAPI-stain. Proportions of whole embryos labeled with chromos ome 1-probe were 17 and 97% at the 3 to 16 and greater than or equal to 32 cell stage, respectively. Of the 93 biopsied embryos analyzed by FISH, 85 e mbryos (91%) could be accurately classified as male or female. Of the 65 bi opsied embryos, 60 embryos (92%) had a clear blastocoele and a inner cell m ass after 48 h of culture in vitro, and these embryos were evaluated as ava ilable embryos. One out of 4 recipient gilts which received sexed embryos a t transfer farrowed 12 piglets of the expected sex. The results of this study demonstrated that porcine embryos at the greater than or equal to 32 cell stage can be sexed within 2 h using the FISH metho d. Moreover further development of the FISH technique could make it an effe ctive tool for the study of early porcine embryos and for the control of po rcine sex. (C) 2000 by Elsevier Science Inc.