System for improving yield of sexed embryos in mammals

• US 6,071,689 A
• Filed: 01/29/1998
• Issued: 06/06/2000
• Est. Priority Date: 12/31/1997
• Status: Expired due to Term

• Current Assignee: XY, LLC (Inguran LLC)
• Original Assignee: XY Incorporated (Inguran LLC)
• Inventors: Schenk, John, Seidel, George, Herickhoff, Lisa
• Primary Examiner(s): Myers, Carla J.
• Application Number:
  US09/015,454
• Time in Patent Office: 859 Days
• Field of Search: 435/6, 435/7.1, 435/7.21, 435/7.2, 435/440, 435/325, 435/2, 800/2, 800/8, 600/33, 600/34, 600/35
• US Class Current: 435/2
• CPC Class Codes: A01K 67/027 New or modified breeds of vertebrates
• CPC Class Codes: A61D 19/02 for artificial insemination
• CPC Class Codes: C12N 15/873 Techniques for producing new embryos, e.g. nuclear transfer, manipulation of totipotent cells or production of chimeric embryos
• CPC Class Codes: C12N 5/0612 sorting of gametes, e.g. according to sex or motility
• CPC Class Codes: G01N 15/149 specially adapted for sorting particles, e.g. by their size or optical properties

Abstract

Improved insemination systems particularly adapted to use for sex-selected sperm sorting include systems which achieve superovulation and then multiple embryo production with sexed embryos. These systems combine with other techniques, including techniques for enhanced sheath fluid and other strategies which minimize stress on the sperm cells, and, potentially, a 2.9 percent sodium citrate sheath solution for bovine species and a hepes bovine gamete media for equine species. Improved collection systems and techniques for the process are described so that commercial application of sperms samples as well as the resulting animals may be achieved.

Claims

1. A method of producing multiple, sexed embryos from a female mammal comprising:

a. creating superovulation in said female mammal to create at least two eggs comprising the step of using an ovulatory pharmaceutical to cause multiple eggs to be produced comprising the steps of;

i) injecting a dosage of follicle stimulating hormone a plurality of times a day;

ii) administering said dosage of follicle stimulating hormone with prostaglandin F-2-alpha;

and wherein said step of injecting said dosage of follicle stimulating hormone a plurality of times a day comprises injecting said follicle stimulating hormone in approximately half day increments at a dosage level of 6, 6, 4, 4, 2, 2, 2, and 2 mg between days 9 and 12 inclusive of the estrus cycle and wherein administering said dosage of follicle stimulating hormone with prostaglandin F-2-alpha comprises supplementing 25 and 12.5 mg of prostaglandin F-2-alpha on the sixth and seventh dosages, respectively, of said follicle stimulating hormone;

b. determining a sex of a sperm cell of a male mammal;

c. sorting according to said sex of said sperm cells;

d. inserting at least a portion of said sorted sperm cells into a uterus of said female mammal after an onset of estrus; and

e. fertilizing a plurality of said eggs to produce at least two sexed embryos of the desired sex from said female mammal.

2. A method of producing multiple, sexed embryos according to claim 1 wherein said step of using a an ovulatory pharmaceutical comprises the step of injecting said ovulatory pharmaceutical in half days increments between any of days 2 and 18 of said estrus cycle.

3. A method of producing multiple, sexed embryos as described in claim 2 wherein injecting said ovulatory pharmaceutical in half day increments comprises injecting at least seven injections and further comprising the step of incorporating an estrus manipulation system at least on about the sixth and seventh injections.

4. A method of producing multiple, sexed embryos as described in claim 3 wherein inserting at least a portion of said sorted sperm cells into said uterus comprises inserting said sperm cells into both uterine horns of said uterus.

5. A method of producing multiple, sexed embryos as described in claim 4 wherein inserting into both uterine horns comprises inserting said sperm cells approximately between 20 to 24 hours inclusive after said onset of said estrus.

6. A method of producing multiple, sexed embryos as described in claim 1 and further comprising the steps of:

a. staining sperm cells of a male mammal;

b. sorting according to said sex of said sperm cells at sperm cell sorting rates above 500 sorts per second; and

c. concentrating said sorted sperm cells.

7. A method of producing multiple, sexed embryos as described in claim 1 wherein inserting at least a portion of said sorted sperm cells comprises using a low dose of said sperm cells.

8. A method of producing multiple, sexed embryos as described in claim 6 wherein inserting at least a portion of said sorted sperm cells comprises using a low dose of said sperm cells.

9. A method of producing a mammal of a desired sex comprising producing said mammal using the processes as described in claims 1.

10. A method of producing a mammal of a desired sex comprising producing a sexed sperm cells mammal using the processes of claim 9 further comprising sorting said cells at sperm cell sorting rates above 500 sorts per second.

11. A method of producing a mammal of a desired sex as described in claim 9 further comprising inseminating a female mammal using a low dose of said sperm cells to produce said mammal of a desired sex.

12. A method of producing multiple, sexed embryos comprising the steps of:

a. creating superovulation in a female mammal to create at least two eggs using an ovulatory pharmaceutical to cause multiple eggs to be produced;

b. chemically coordinating a sheath fluid to create a sheath fluid environment for sperm cells of a male mammal which is coordinated with both a pre-sort and a post-sort cell fluid environment comprising establishing a sheath fluid which contains about 2.9% sodium citrate;

c. determining a sex of said sperm cells;

d. sorting according to said sex of said sperm cells;

e. inserting at least a portion of said sorted sperm cells into a uterus of said female mammal after an onset of estrus; and

f. fertilizing a plurality of said eggs to produce at least two sexed embryos of the desired sex from said female mammal.

13. A method of producing multiple, sexed embryos as described in claim 12 wherein chemically coordinating a sheath fluid to create a sheath fluid environment for said cells which is coordinated with both a pre-sort and a post-sort cell fluid environment comprises establishing a sheath fluid which contains a HEPES buffered medium.

14. A method of producing multiple, sexed embryos as described in claim 13 further comprising collecting cells of said desired sex and cushioning said cells from impact with a collection container which has a wide opening.

15. A method of producing multiple, sexed embryos as described in claim 12 and further comprising the steps of:

a. staining sperm cells of a male mammal;

b. sorting according to said sex of said sperm cells at sperm cell sorting rates above 500 sorts per second; and

c. concentrating said sorted sperm cells.

16. A method of producing multiple, sexed embryos as described in claim 12 wherein inserting at least a portion of said sorted sperm cells comprises using a low dose of said sperm cells.

17. A method of producing multiple, sexed embryos as described in claim 15 wherein inserting at least a portion of said sorted sperm cells comprises using a low dose of said sperm cells.

18. A method of producing a mammal of a desired sex comprising producing said mammal using the processes as described in claim 12.

19. A method of producing a mammal of a desired sex comprising producing said mammal using the processes as described in claim 13.

20. A method of producing a mammal of a desired sex comprising producing said mammal using the processes as described in claim 16.