Method to preselect the sex of offspring
AssigneeThe United States of America as represented by the Secretary of Agriculture
InventorLawrence A. Johnson
Life of this patent
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Abstract
Intact X and Y chromosome-bearing sperm populations of rabbits and swine were separated according to DNA content using a flow cytometer/cell sorter. Sperm viability was maintained by special staining techniques and by sorting and collecting the sperm in nutrient media. The sorted sperm were surgically inseminated into the uteri of rabbits or swine. Of the offspring born from does inseminated with the sorted population of X-bearing sperm, 94% were females. Of offspring born from does inseminated with sorted Y-bearing sperm from the same ejaculate, 81% were males.
Claims26
A method for sorting intact, viable, mammalian sperm into X- and Y-chromosome-bearing populations based on DNA content, the method comprising:
- a) staining intact, viable sperm collected from a male mammal with a fluorescent dye capable of selectively staining DNA in living cells by incubating the sperm with the dye at a temperature in the range of about 30.degree.-39.degree. C. for a period of time sufficiently long for staining to take place uniformly but sufficiently short to preserve viability of the sperm;
- b) passing the sperm into an electrically conductive and isotonic viability-supporting sheath fluid to form a suspension of sperm which are caused to flow singly in a stream of sheath fluid;
- c) passing the sheath fluid containing the sperm before an excitation light source causing the stained DNA to fluoresce;
- d) passing the sheath fluid containing the sperm through both a means for detecting the fluorescence of the stained DNA and also a cell sorting means, the means for detecting fluorescence having at least two detectors arranged such that a first detector determines the orientation of sperm on the basis of magnitude of fluorescence and controls a second detector to measure the DNA content of sperm on the basis of magnitude of fluorescence of those sperm that have been determined to be in a preselected orientation;
- e) selecting by said cell sorting means the sperm having a DNA content corresponding to a desired chromosome which will produce a desired gender of offspring, and separating the selected sperm from nonselected sperm;
- and
- f) collecting the selected sperm in a viability-supporting collecting fluid.
A method to preselect the sex of mammalian offspring comprising:
- a) staining intact, viable sperm collected from a male mammal with a fluorescent dye capable of selectively staining DNA in living cells by incubating sperm with the dye at a temperature in the range of about 30.degree.-39.degree. C. for a period of time sufficiently long for staining to take place uniformly but sufficiently short to preserve viability of the sperm;
- b) passing the sperm into an electrically conductive and isotonic viability-supporting sheath fluid to form a suspension of sperm which are caused to flow singly in a stream of sheath fluid;
- c) passing the sheath fluid containing the sperm before an excitation light source causing the stained DNA to fluoresce;
- d) passing the sheath fluid containing the sperm through both a means for detecting the fluorescence of the stained DNA and also a cell sorting means to measure the DNA content of the sperm on the basis of magnitude of fluorescence of the sperm;
- e) selecting by said cell sorting means the sperm having a DNA content corresponding to a desired chromosome which will produce the desired gender of offspring, and separating the selected sperm from nonselected sperm;
- and
- f) collecting the selected sperm in a viability-supporting collecting fluid.
A method for preparing intact, viable, mammalian sperm for sorting into X- and Y-chromosome-bearing populations based on DNA content, the method comprising staining intact, viable sperm collected from a male mammal with a fluorescent dye capable of selectively staining DNA in living cells by incubating the sperm with the dye at a temperature in the range of about 30.degree.-39.degree. C. for a period of time sufficiently long for staining to take place uniformly but sufficiently short to preserve viability of the sperm.
Dependencies are inferred from the claim text.