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Studies on sorted x and y enriched populations of bull spermatozoa

Published online by Cambridge University Press:  24 November 2017

N.G.A. Miller ,
E.A. Howes ,
D.G. Cran  and
L.A. Johnson
N.G.A. Miller
Affiliation:
I.A.P.G.R. Babraham Hall, Babraham, Cambs. CB2 4AT
E.A. Howes
Affiliation:
I.A.P.G.R. Babraham Hall, Babraham, Cambs. CB2 4AT
D.G. Cran
Affiliation:
A.B.C. Ltd., Cambridge CB3 OJQ
L.A. Johnson
Affiliation:
U.S.D.A., Beltsville, Maryland 20705, USA.
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Extract

There have been many attempts to separate X- and Y-chromosome bearing spermatozoa on the basis of surface charge or density (Schröder, 1941; Bhattacharya et at., 1966; Engelman et al., 1988; Blottner et al., 1992). These have not met with universally accepted success. More recently, sorting has been achieved using the flow cytometer (FACS) to discriminate between the very small differences (3-4%) in DNA content of X- and Y-bearing spermatozoa (Johnson et al., 1989; Cran et al., 1993). However, the spermatozoa must be ‘flat headed’ e.g. rabbit, chinchilla, horse, pig, cattle and sheep.

The flow cytometer (fitted with a U.V. laser) was adapted to accept a bevelled inlet tube. This gave us a ‘ribbon-like’ profile of the sample stream which forced the spermatozoa into a specific orientation.

Semen was collected from healthy bulls of proven fertility and within 1/2 hr. was diluted to 1 x 107 per ml. and stained with Hoechst 33342 (5-10/μm) for 1 hr. at 35°C. Only those spermatozoa emerging from the 70μ nozzle that were edge-on to the 90° detector were selected as X or Y by the 0° detector. Gated intact spermatozoa were positioned into individual drops and then charged positive or negative for later deflection and collection into tubes containing egg yolk medium.

Type
The Application of Molecular Biology to Animal Science
Information
Proceedings of the British Society of Animal Production (1972) , Volume 1993: Winter meeting , March 1993 , pp. 156
Copyright
Copyright © The British Society of Animal Production 1993

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References

Bhattacharya, B.C., Bangham, A.D., Cro, R.J. and Keynes, R.D. (1966) An attempt to predetermine the sex of calves by artificial insemination with spermatozoa separated by sedimentation. Nature 211, 863.CrossRefGoogle ScholarPubMed
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Cran, D.G., Johnson, L.A., Miller, N.G.A., Cochrane, D. and Polge, C. (1993) Production of bovine calves following separation of X- and Y-chromosome bearing sperm and in vitro fertilization. The Veterinary Record 132.Google Scholar
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