돼지정액의 동결에 관한 연구 II. 동결한 돼지정액의 체내, 체외수정능력

Studies on the Freezing of Boar Semen II . In Vitro and In Vivo Fertilizing Capacity of Frozen Boar Spermatozoa

  • 발행 : 2005.04.01

초록

본 실험은 LEY에 다양한 당류를 첨가한 용액으로 동결보존한 돼지정자의 응해후 생존율과 정상 첨체율, 체외성숙한 난자와의 수정능력과 배 발달 능력 및 인공수정을 통한 수태율과 산자수에 미치는 효과를 조사하기 위해 실시하였다. 융해후의 정상 첨체율은 glucose 농도 증가와 함께 증가되었으나, 생존율에 있어서는 효과가 인정되지 않았다. LEY에 fructose를 단독 또는 glucose와 함께 첨가하면 융해후의 생존율이 유의하게 증가되었으며 (p<0.05), 정상 첨체율에 있어서는 LEY 보존액에 fructose와 glucose를 첨가한 구가 $81.4{\pm}2.3\%$로 control의 $41.6{\pm}0.6\%$에 비하여 유의하게 높았다(p<0.001). 체외성숙 난자와의 수정율, 분할율 및 배반포 발생율은 $70.8\~80.7\%$, $44.6\~45.7\%$$13.6\~16.0\%$로 glycerol과 fructose 농도 및 정자농도 간에 차이가 인정되지 않았다. 1회 발정당 2회의 인공수정을 하였을 때, 수태율은 $83.3{\pm}0.1\%$, 산자수는 $9.4{\pm}1.7\~10.4{\pm}0.7$두로서 SGI사의 동결정액의 $50.0{\pm}0.1\%(p<0.05)$과 $8.0{\pm}1.1$두에 비하여 높았다.

This experiment was carried out to investigate the effects of saccharide in the lactose-egg yolk(LEY) extender for freezing of boar semen on the viability, normal acrosome, fertilizable of in vitro or in vivo oocyte after thawed. Normal acrosome post-thawed spermatozoa was higher when increasing of glucose concentration in LEY extender with 3 or $4\%$ glycerol, but viability was not significant. Viability of the post-thawed spermatozoa was higher when fructose or fructose and glucose were added to LEY extender with $3\%$ glycerol than glucose and sucrose or fructose, glucose and sucrose(P<0.05). Rate of normal acrosome of post thawed spermatozoa was higher when both fructose and glucose$(81.4{\pm}2.3\%)$ were added to the LEY extender than saccharide not added$(41.6\pm0.6\%)$ to it(P<0.001). The percentage of fertilization, cleavage and development to blastocyst of oocytes fertilized with post-thawed spermatozoa from freezing by LEY extender were $70.8\~80.7\%$, $44.6\~45.7$ and $13.6\~16.0\%$, respectively. Conception rate by artificial insemination with frozen boa. semen was higher$(83.1{\pm}0.3\%)$ than commercial frozen semen from SGI company$(50.0{\pm}0.1\%,\;P<0.05)$, but litter size were no significant differences between frozen by LEY extender$(9.4{\pm}1.7\~10.4{\pm}0.7head/sow)$ and SGI semen$(8.0{\pm}1.1 head/sow)$.

키워드

참고문헌

  1. Abeydeera LR and Day BN. 1997a. In vitro penetration of pig oocytes in a modified Tris-buffered medium: Effect of BSA, caffeine and calcium. Theiogenology, 48:537-544 https://doi.org/10.1016/S0093-691X(97)00270-7
  2. Almlid T, Clarke VG, Pursel VG and Johnson LA. 1989. Effectiveness of in vitro methods for predicting in vitro fertilizing capacity of boar spermatozoa cryopreserved with 2% or 4% glycerol. Zuchthygiene, 24:8-15 https://doi.org/10.1111/j.1439-0531.1989.tb00655.x
  3. Almlid T and Johnson LA 1988. Effects of glycerol concentration, equilibration time and temperature of glycerol addition on post-thaw viability of boar spermatozoa in straws. J. Anim. Sci., 66:2899-2905 https://doi.org/10.2527/jas1988.66112899x
  4. Barth AD. 1992. The relationship between sperm abnormalities and fertility. In: Proc 14th Tech Conf Artif Insem and Reprod. Nat. Assoc. Anim. Breeders, Columbia, Missouri. p. 47
  5. Bavister BD. 1990. Test of sperm fertilizing ability. In: Asch RH, Balmaceda JP, Johnson I(eds), Gamete Physiology. Serono Symposia, USA pp. 77-705
  6. Eriksson BM and Rodriguez-Martinez H. 2000. Export of frozen boar semen in a new flat package. 4th Int. Conf. Boar semen preservation, p. 244
  7. Fiser PS and Fairfull RW. 1990. Combined effect of glycerol concentration and cooling velocity on motility and acrosomal integrity of boar spermatozoa frozen in 0.5 ml. straws. Mol. Reprod. Dev., 25:123-129 https://doi.org/10.1002/mrd.1080250204
  8. Funahashi H and Day BN. 1993. Effect of the duration of exposure to supplemental hormones on cytoplasmid maturation of pig oocytes in vitro. J. Reprod. Fertil., 98:179-185 https://doi.org/10.1530/jrf.0.0980179
  9. Garner DL and Johnson LA. 1995. Viability assessment of mammalian sperm using SYBR-14 and propidium iodide. Biol, Reprod., 53:276-284 https://doi.org/10.1095/biolreprod53.2.276
  10. Johnson LA. 1985. Fertility results using frozen boar spermatozoa: 1970 to 1985. 1st Int. Conf. on Deep Freezing of Boar Semen. Swedish Univ. Agric. Sciences, Uppsala. pp. 199-222
  11. Johnson LA. 1992. Gender preselection in swine: Altered sex ratios in offspring following surgical insemination of flow sorted X- and Y-bearing sperm. Reprod. Dom. Anim., 26:309-314 https://doi.org/10.1111/j.1439-0531.1991.tb01546.x
  12. Johnson LA, Weitze KF, Fiser P and Maxwell WMC. 2000. Storage of boar semen. Anim. Reprod. Sci., 62:143-172 https://doi.org/10.1016/S0378-4320(00)00157-3
  13. Maxwell WMC and Johnson LA. 1997b. Membrane status of boar spermatozoa after cooling or cryopreservation. Theriogenology, 48:209-219 https://doi.org/10.1016/S0093-691X(97)84068-X
  14. Nagai T, Takahashi H, Masuda Y, Shioya M, Kuwayama M, Fukushima M, Iwasaki Sand Hanada A. 1988. In vitro fertilization of pig oocytes by frozen boar spermatozoa. J. Reprod. Fertil., 84:585-591 https://doi.org/10.1530/jrf.0.0840585
  15. Obando H, Tamayo T and Alvaro C. 1984. Evaluation of some factors affecting swine spermatozoa during freezing. X th Intern. Congo Anim. Reprod. AI. Ubana, II, 193
  16. Paquignon M. 1985. Freezing and thawing extenders for boar spermatozoa. In: Deep freezing of boar semen. Proc. 1 st Int. Con. Deep Freez. Boar Semen. Uppsala. p:129-145. Eds. Johnson, L.A. and Larsson, K. Swedish University of Agricultural Sciences, Uppsala
  17. Petters RM and Wells KD. 1993. Culture of pig embryos. J. Reprod. Fertil., 48:(Suppl)61-73
  18. Rath D, Niemann H and Tao T. 1995. In vitro maturation of porcine oocytes in follicular fluid with subsequent effects on fertilization and embryo yield in vitro. Theriogenology, 44:529-538 https://doi.org/10.1016/0093-691X(95)00224-V
  19. Reed HCB. 1985. Current use of frozen boar semen future need of frozen boar semen. In: Deep freezing of boar semen. Proc. 1 st Int. Con. Deep Freez. Boar semen. Uppsala. p:225-237. Eds. Johnson, L.A. and Larsson, K. Swedish University of Agricultural Sciences, Uppsala
  20. Rodriguez-Martinez H, Eriksson Band Lundeheim I. 1996. Freezing boar semen in flat plastic bags. Membrane integrity and fertility. In: Rath, D., Johnson, L.A. and Weitze, K.F.(Eds). Boar Semen Preservation Ill. Proc. 3rd Int. Conf. Deep Freezing Boar Semen. Reprod. Dom. Anim. 31 Blackwell, Berlin. pp. 161-168 (Suppl. 1)
  21. Salamon S, Wilmut I and Polge C. 1973a. Deep freezing of boar semen. I. Effects of diluent composition, protective agents and method of thawing on survival of spermatozoa. Aust. J. Biol. Sci., 26:219-230
  22. Visser D and Salamon S. 1973. Fertility of ram spermatozoa frozen in a trisbased diluent. Aust. J. Biol. Sci., 26:513-516 https://doi.org/10.1071/BI9730513
  23. Westendorf P, Richter Land Treu H. 1975. Zur Tierfgefrierung von Embersperma: Labor und Besamungsergebnisse mit dem Hulsenberger Pailleten Verfahren. Dtch. tierarztl. Wschr, 82: 261-267
  24. Wilmut I and Polge C. 1977a. The low temperature preservation of boar spermatozoa. I. The motility and morpology of boar spermatozoa frozen and thawed in the presence of permeating protective agents. Cryobiology, 14:471-478 https://doi.org/10.1016/0011-2240(77)90009-8
  25. Wilmut I and Polge C. 1977b. The low temperature preservation of boar spermatozoa. 2. The motility and morphology of boar spermatozoa frozen and thawed in diluent which contained only sugar and egg-yolk. Cryobiology, 14:479-482 https://doi.org/10.1016/0011-2240(77)90010-4
  26. Weitze KF, Rath D and Baron G. 1987. Neue Aspekte der Tiefgefrierkonservierung von Eber-sperma in Plastikrohren. Dtsch. tierrztl, Wschr., 94:485-488
  27. Yoshida M, Ishizaki Y and Kawagishi H. 1990. Blastocyst formation by pig embryos resulting from in vitro fertilization of oocytes matured in vitro. J. Reprod. Fertil., 88:1-8 https://doi.org/10.1530/jrf.0.0880001
  28. 川倉一彦, 副島昭彦, 田博司. 1984. Straw법에 의한 돼지정액의 동결보존. 일본가축인공수정연구지. 6:61-64