Production of Transgenic Rabbits Using Centrifuged Pronuclear Zygotes

Journal of Veterinary Medical Science

Ken KIMURA³  Ri-ichi TAKAHASHI¹  Shinichi HOCHI³  Masao HIRAO²  Masatsugu UEDA¹  Masumi HIRABAYASHI¹  Kazuo HIRASAWA² 

[1] YS New Technology Institute Inc.,Ishibashi-Machi,Shimotsuga-Gun,Tochigi 329-0512,Japan;Ina Research Laboratory,Kitayama Labes Co.Ltd.,Ina,Nagano 396-0021,Japan;Faculty of Textile Science and Technology,Shinshu University,Ueda,Nagano 386-8567,Japan

关键词: centrifugation;    cytoplasmic lipid;    hGH;    pronucleus;    transgenic rabbit;   

DOI  :  10.1292/jvms.62.1047

学科分类：兽医学

来源: Japanese Society of Veterinary Science

【 摘 要 】

References(35)Cited-By(6)Superovulation of female rabbits was induced by subcutaneous injection(s)of porcine FSH.Zygotes were recovered 17 to 19 hr after hCG injection and were classified into two categories under a microscope equipped with Nomarski interference−contrast optics at ×200 magnification:(A)zygotes with clearly visible pronuclei, or(B)zygotes with visualized pronuclei after 10 min centrifugation at 12, 000×g.No significant difference between strains was found in the proportion of category−A zygotes(JW 72.6% vs NZW 79.3%).Pronuclei of category−A zygotes were located in the center of the cytoplasm, and the pronuclei of category−B zygotes were slightly moved by centrifugation toward the mass of cytoplasmic lipid droplets.Exogenous DNA solution(5 μg/ml of fusion gene composed of bovine αS1−casein promoter and human growth hormone structural gene)was microinjected into the pronucleus of the JW zygotes.The pronucleus of category−A zygotes with a mean volume of 7.4 pl swelled up to 16.6 pl(132% increase), while that of category−B zygotes with a mean volume of 6.1 pl swelled up to 15.9 pl(148% increase).Nevertheless, similar proportions of category−A and category−B zygotes developed into offspring after transfer to recipient females(11.1 and 11.2%, respectively).The efficiency to produce hGH−carrying transgenic rabbits was 0.9%(2/235)from category−A zygotes and 0.5%(1/215)from category−B zygotes(P>0.05).To date, transgenic rabbits have been produced without centrifugation of pronuclear zygotes.However approximately 25% of fertilized rabbit zygotes can be used for DNA microinjection after they have been centrifuged to visualize their pronuclei.

【 授权许可】

Unknown