외래유전자 도입정자를 이용한 돼지 체외성숙 난포란의 Intracytoplasmic Sperm Injection (ICSI) 후 후기 배로의 발달율과 외래유전자의 발현에 관한 연구

서지정보

· 발행기관 : 한국수정란이식학회
· 수록지 정보 : Journal of Embryo Transfer / 17권 / 1호
· 저자명 : 정기화, 조성근

초록

본 연구는 sperm-mediated gene transfer를 이용하여 ICSI에 의한 형질전환동물 생산의 기초자료로서 활용하기 위해, ICSI에 사용될 정자의 조건과, 그에 따라 적합한 돼지 정자와 외래유전자의 전처리 및 ICSI를 통한 수정을 및 후기 배로의 발달율과 외래유전자의 발현 여부를 조사하여 다음과 같은 결과를 얻었다. ICSI에 이용될 정자의 조건에 따라 정소상체미부정자, 사출정자 및 동결정자를 이용하여 ICSI후 수정율은 각각 72.3

영어초록

The main goal of this study was to produce transgenic porcine embryos by direct injection of sperm-mediated exogenous DNA. Spermatozoa (610 sperms of final concentration) were mixed with pcDNA LAC Z (20 ng/l) and subjected into electroporation (300~750 volts, 25 F, 0.4 cm electrode). After sperm injection, the oocytes were activated electrically (1.7 KV/cm, 30sec, single pulse) in 0.3 M mannitol solution or not. The sperm injected eggs were cultured in NCSU 23 medium (0.4% BSA) at 39, 5% in air fur 144 h. The rates of cleavage and development into blastocyst stage in activation group were significantly higher than those of non-activation group (79.6% and 24.1% vs. 46.3% and 14.4%, respectively, p<0.05). Control oocytes and shame injection were developed to blastocysts low (2.5%). Sixty five (27.1%) out of 240 embryos observed in activation and non-activation groups were showed positive by X-gal staining. However, all embryos in both groups were expressed partial or mosaic pattern. These results suggested that electrical stimulation far oocytes activation after sperm injection enhances the incidence of both fertilization and development fellowing sperm injection in the pig. Our study also suggested that sperm-mediated transfer of exogenous DNA by ICSI would be used as a valuable tool for the production of transgenic porcine embryos.

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