Journal of Aquaculture (한국양식학회지)

The Korean Society of Fisheries and Aquatic Science (한국양식학회)
권호 표지

Intraspecific Androgenesis in Mud Loach (Misgurnus mizolepis): II. Diploid Restoration and Viability Assessment

미꾸라지(Misgurnus mizolepis)의 웅성발생성 처녀생식: II. 웅성발생성 이배체 유도 및 생존능력 평가

  • Nam, Yoon-Kwon (Department of Aquaculture & Institute of Marine Living Modified Organisms (iMLMO), Pukyong National University) ;
  • Bang, In-Chul (Department of Marine Biotechnology, Soonchunhyang University) ;
  • Noh, Choong-Hwan (Marine Resources and Research Department, Korea Ocean Research and Development Institute) ;
  • Cho, Young-Sun (Department of Aquaculture & Institute of Marine Living Modified Organisms (iMLMO), Pukyong National University) ;
  • Kim, Dong-Soo (Department of Aquaculture & Institute of Marine Living Modified Organisms (iMLMO), Pukyong National University)
  • 남윤권 (부경대학교 양식학과/해양수산형질전환생물 연구소) ;
  • 방인철 (순천향대학교 해양생명공학과) ;
  • 노충환 (한국해양연구원 생물자원본부) ;
  • 조영선 (부경대학교 양식학과/해양수산형질전환생물 연구소) ;
  • 김동수 (부경대학교 양식학과/해양수산형질전환생물 연구소)
  • Published : 2006.11.25
Download PDF
⟨ Previous Next ⟩

Abstract

Intraspecific diploid androgenesis was achieved in mud loach (Misgurnus mizolepis) by the inhibition of the first mitotic division using combined thermal treatment. A combined thermal treatment (heat shock at $40.5\;^{\circ}C$ for 120 sec followed by cold treatment at $1\;^{\circ}C$ for 45 min) applied to the 1st metaphase of cell division (28 min post insemination at $25\;^{\circ}C$) successfully recovered viable androgenetic diploidy. Mean hatching success of the androgenetic diploid group was 29.6%, and the average yield out of total eggs taken was about 7% assessed at 1 week of age. However, relatively large variations in the yield of diploid androgenesis were observed among different egg batches used as cytoplasmic donors. Successful diploidization was confirmed by flow cytometric analysis, and parthenogenic reproduction in a paternal exclusive manner was verified with transgene dosage. Significant mortality was found in most androgenetic groups especially from hatch to 1 month of age, although such mortality was stabilized later.

우리나라 주요 담수어종인 미꾸라지(Misgurnus mizolepis)를 어류 염색체 조작 모델로 개발하기 위한 연구의 일환으로 동일 종내 웅성발생성 이배체 유도 조건을 개발하고 유도된 웅성발생성 이배체의 생존능력을 평가하였다. 자외선(UV; $10,800\;ergs/mm^2$)을 이용하여 난자의 유전물질을 불활성화시키고 인공수정 28분 후(제1난할 중기)에 온도처리(고온 자극 $40.5\;^{\circ}C$ 120초 후 저온 자극 $1\;^{\circ}C$ 45분)를 통해 제1난할을 억제하였다. 본 온도처리를 통해 생존력 있는 웅성발생성 이배체들이 유도되었고(평균 부화율, 26.9%), 부화 후 1주일째를 기준으로 평균 웅성발생성 이배체 수율은 최초 처리난의 7%내외로 나타났다. 그러나 웅성발생성 이배체 유도 효율은 cytoplasmic donor의 질에 크게 영향을 받음이 관찰되어 사용한 암컷 친어별로 큰 편차가 관찰되었다. Flow cytometry 분석에 의해 생존력 있는 웅성발생성 개체들의 이배체 복원이 확인되었으며 아울러 형질전환 표지를 이용하여 동형접합성 개체 유도의 가능성이 확인되었다. 웅성발생성 이배체들은 일반대조군 이배체에 비해 낮은 생존능력을 보였으며 특히 부화 후 1개월까지 유의적으로 낮은 생존율을 나타내었다. 그러나 후기 생존능력은 일반 이배체군과 유사하였다.

Keywords

References

  1. Arai, K., 2001. Genetic improvement of aquaculture finfish species by chromosome manipulation techniques in Japan. Aquaculture, 197, 205-228 https://doi.org/10.1016/S0044-8486(01)00588-9
  2. Babiak, I., S. Dobosz, H. Kuzminski, K. Goryczko, S. Ciesielski, P. Brzuzan, B. Urbanyi, A. Horvath, F. Lahnsteiner and J. Piironen, 2002b. Failure of interspecies androgenesis in salmonids. J. Fish. Biol., 61, 432-447 https://doi.org/10.1111/j.1095-8649.2002.tb01575.x
  3. Babiak, I., S. Dobosz, K. Goryczko, H. Kuzminski, P. Brzuzan and S. Ciesielski, 2002a. Androgenesis in rainbow trout using cryopreserved spermatozoa: the effect of processing and biological factors. Theriogenology, 57, 1229-1249 https://doi.org/10.1016/S0093-691X(02)00631-3
  4. Bercsenyi, M., I. Magyary, B. Urbanyi, L. Orban and L. Horvath, 1998. Hatching out goldfish from common carp eggs: interspecific androgenesis between two cyprinid species. Genome, 41, 573-579 https://doi.org/10.1139/gen-41-4-573
  5. Bongers, A. B. J., E. P. C. in't Veld, K. Avo-Hashema, I. M. Bremmer, E. H. Eding, J. Komen and C. J. J. Richter, 1994. Androgenesis in common carp (Cyprinus carpio L.) using UV irradiation in a synthetic ovarian fluid and heat shocks. Aquaculture, 122, 119-132 https://doi.org/10.1016/0044-8486(94)90504-5
  6. Bongers, A. B. J., J. B. Abraca, Z. Doulabi, E. H. Eding, J. Komen and C. J. J. Richter, 1995. Maternal influence on development of androgenetic clones of common carp, Cyprinus carpio L. Aquaculture, 137, 139-147 https://doi.org/10.1016/0044-8486(96)83544-7
  7. Cho, Y. S., B. N. Choi, E.-M. Ha, K. H. Kim, S. K. Kim, D. S. Kim and Y. K. Nam, 2005. Shark, Scyliorhinus torazame metallothionein: cDNA cloning, genomic sequence and expression analysis. Mar. Biotechnol., 7, 350-362 https://doi.org/10.1007/s10126-004-0043-y
  8. Corley-Smith, G. E., C. J. Lim and B. P. Brandhorst, 1996. Production of androgenetic zebrafish (Danio rerio). Genetics, 142, 1265-1276
  9. David, C. J. and T. J. Pandian, 2006. Cadaveric sperm induces intergeneric androgenesis in the fish, Hemigrammus caudovittatus. Theriogenology, 65, 1048-1070 https://doi.org/10.1016/j.theriogenology.2005.07.014
  10. Kim, D. S., J.-Y. Jo, and T.-Y. Lee, 1994. Induction of triploidy in mud loach (Misgurnus mizolepis) and its effect on gonad development and growth. Aquaculture, 120, 263- 270 https://doi.org/10.1016/0044-8486(94)90083-3
  11. Kirankumar S. and T. J. Pandian, 2004. Interspecific androgenetic restoration of rosy barb using cadaveric sperm. Genome, 47, 66-73 https://doi.org/10.1139/g03-104
  12. Kirankumar, S. and T. J. Pandian, 2003. Production of androgenetic tiger barb Puntius tetrazona. Aquaculture, 228, 37-51 https://doi.org/10.1016/S0044-8486(03)00132-7
  13. Lin, F. and K. Dabrowski, 1998. Androgenesis and homozygous gynogenesis in muskellunge (Esos masquinongy): evaluation using flow cytometry. Mol. Reprod. Dev., 49, 10-18 https://doi.org/10.1002/(SICI)1098-2795(199801)49:1<10::AID-MRD2>3.0.CO;2-R
  14. Maremgpmo, M. G. and Y. Onoue, 1998. Ultraviolet-induced androgenesis in nile tilapia, Oreochromis niloticus (L.), and hybrid nile x blue tilapia, O. aureus (Steindachner). Aquaculture Res., 29, 359-366
  15. Nam, Y. K., C. H. Noh and D. S. Kim, 1999. Transmission and expression of an integrated reporter construct in three generations of transgenic mud loach (Misgurnus mizolepis). Aquaculture, 172, 229-245 https://doi.org/10.1016/S0044-8486(98)00433-5
  16. Nam, Y. K., C. H. Noh and D. S. Kim, 2006. Intraspecies androgenesis in mud loach (Misgurnus mizolepis): I. Optimization of the egg inactivation and haploid androgenesis using transgene marker. J. Aquacult., 19 (in press)
  17. Nam, Y. K., G. C. Choi and D. S. Kim, 2004. An efficient method for blocking the 1st mitotic cleavage of fish zygote using combined thermal treatment, exemplified by mud loach (Misgurnus mizolepis). Theriogenology, 61, 933-945 https://doi.org/10.1016/S0093-691X(03)00258-9
  18. Nam, Y. K., Y. S. Cho, H. J. Cho and D. S. Kim, 2002. Accelerated growth performance and stable germ-line transmission in androgenetically derived homozygous transgenic mud loach, Misgurnus mizolepis. Aquaculture, 209, 257-270 https://doi.org/10.1016/S0044-8486(01)00730-X
  19. Nam, Y. K., Y. S. Cho, Y. J. Chang, J.-Y. Jo and D. S. Kim, 2000. Generation of transgenic homozygous line carrying the CAT gene in mud loach Misgurnus mizolepis. Fish. Sci., 66, 58-62 https://doi.org/10.1046/j.1444-2906.2000.00008.x
  20. Pandian, T.J. and R. Koteeswaran, 1998. Ploidy induction and sex control in fish. Hydrobiologia, 384, 167-243 https://doi.org/10.1023/A:1003332526659
  21. Paschos, I., L. Natsis, C. Nathanailides, I. Kagalou and E. Kolettas, 2001. Induction of gynogenesis and androgenesis in goldfish Carassius auratus (var. oranda). Reprod. Dom. Anim., 36, 195-198 https://doi.org/10.1046/j.1439-0531.2001.d01-36.x
  22. Tanck, M. W. A. P. Palstra, M. van der Weerd, C. P. Leffering, J. J. van der Poel, H. Bovenhuis and J. Komen, 2001. Segregation of microsatellite alleles and residual heterozygosity at single loci in homozygous androgenetic common carp (Cyprinus carpio L.). Genome, 44, 743-751
  23. Thorgaard, G. H., 1986. Ploidy manipulation and performance. Aquaculture, 57, 57-64 https://doi.org/10.1016/0044-8486(86)90180-8
  24. Zhang, H., B., Robison, G. H. Thorgaard and S. S. Ristow, 2000. Cloning, mapping and genomic organization of a fish C-type lectin gene from homozygous clones of rainbow trout (Oncorhynchus mykiss). Biochim. Biophys. Acta, 1494, 14-22 https://doi.org/10.1016/S0167-4781(00)00198-6