An X-Y paint set and sperm FISH protocol that can be used for validation of cattle sperm separation procedures.
X and Y chromosome paints were developed from sorted yak chromosomes for sexing cattle spermatozoa. Clear hybridization signals were obtained for every spermatozoon using a modified sperm decondensation protocol and fluorescence in situ hybridization (FISH). The procedure was evaluated using the established Beltsville sperm sexing technology, which separates spermatozoa by flow cytometry into X- and Y-bearing fractions. Close agreement was found between the assessment of sperm separation by flow cytometry and by FISH with the X-Y paint set. The FISH method is a simple, reliable and robust procedure for assessing the effectiveness of separation of X and Y spermatozoa.
Rens
W
W
Centre for Veterinary Science, Department of Clinical Veterinary Medicine, Veterinary Cytogenetics Group, University of Cambridge, Madingley Road, Cambridge CB3 0ES, UK. wr203@mole.bio.cam.ac.uk
Yang
F
F
Welch
G
G
Revell
S
S
O'Brien
P C
PC
Solanky
N
N
Johnson
L A
LA
Ferguson Smith
M A
MA
D000818:Animals; D002417:Cattle; D002469:Cell Separation / Q000379:methods*; D002478:Cells, Cultured; D020223:Chromosome Painting; D004247:DNA / Q000032:analysis / Q000737:chemistry; D005434:Flow Cytometry; D017404:In Situ Hybridization, Fluorescence; D008297:Male; D012732:Sex Determination Analysis; D013094:Spermatozoa / Q000166:cytology* / Q000648:ultrastructure; D014960:X Chromosome; D014998:Y Chromosome
D004247:DNA