• Asian-Australasian Journal of Animal Sciences
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• 제6권4호
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• pp.541-547
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• 1993

A phage clone containing the partial ${\alpha}_{S1}$-casein gene was isolated from a bovine genomic library by using mixed probes of ovine ${\alpha}_{S1}$-, ${\beta}$- and ${\kappa}$-casein cDNAs. Restriction enzyme mapping analysis for 14.6 kb revealed that the map was in conflict with the report of Meade et al. (1990), especially in the 3'-end fragment. Sequence analysis of 12.6 kb revealed a high AT/GC ratio (1.64); we have identified eight exon sequences according to the bovine ${\alpha}_{S1}$-casein cDNA sequence. The same exon/intron splice junction sequence was observed between these exons. We suggest that the bovine ${\alpha}_{S1}$-casein gene night contain a minimum of 18 exons and the full length is approximately 18-19 kb.

• 한국수정란이식학회:학술대회논문집
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• 한국수정란이식학회 2002년도 국제심포지엄
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• pp.74-74
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• 2002

Production of u 1-antitrypsin ($\alpha$AT) in transgenic cows has a great value in the field of medicine. The present study was conducted to determine the effect of chemically defined KSOM media on in vitro development of bovine transgenic nuclear transfer (NT) embryos. An expression plasmid for human $\alpha$AT was constructed by inserting a bovine beta-casein promoter, a green fluorescent protein (GFP) marker gene, and a human $\alpha$AT target gene into a pcDNA3 plasmid. Cumulus cells as donor nuclei in NT were collected from a Holstein cow and transfected by lipid-mediated method using FuGene6 (Roche Molecular Biochemicals, USA) as reagent. GFP expressed cumulus cells were introduced into recipient oocytes under DIC microscopy equipped with FITC filter set. After electrical fusion and chemical activation, reconstructed embryos were cultured in 1) SOF + 0.8% BSA, 2) KSOM + 0.8% BSA, 3) KSOM + 10% FBS and 4) KSOM +0.01% PVA for 192 h at 39$^{\circ}C$ with 5% $CO_2$, 5% $O_2$ and 90% $N_2$in humidified condition. The development of the embryos was recorded and the GFP expression in blastocyst was determined under FITC filter. The average fusion rate was 73.8% (251/340; n=8). The development rates to 2-4 cells, morula, blastocysts and expression rates in blastocysts varied from 70.3 to 76.5%, 30.2 to 33.8%, 25.4 to 33.8% and 11.8 to 15.6%, respectively. The difference in development and expression rates of embryos among 4 culture groups was not significant (P>0.05). This study indicates that chemically defined KSOM medium is also able to support development of bovine transgenic NT embryos at similar rate of SOF or KSOM supplemented with BSA or serum.

• 한국가축번식학회지
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• 제21권1호
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• pp.1-7
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• 1997

형질 전환동물의 유선을 이용한 단백질의 생산이 보편화되고 있지만 원하는 단백질이 만들어지기 까지 많은 시간과 노력이 필요하며 기술적인 어려움이 항시 따른다. 그래서 보다 쉽게 재조합된 유전자의 발현 정도를 시험하는 in vitro에서의 시험방법의 개발은 중요한 의미가 있다고 하겠다. 따라서 본 연구에서는 인간의 유방암을 가진 환자의 유선에서 유래된 MCF7 cell line을 이용하여 유선 특징의 유전자 발현을 위한 세포 배양 모델을 개발하고자 하였다. 우선 소의 카제인의 cDNA를 MMTV-LTR의 통제하에 clone하였으며, 이것을 CaPO4의 침전법으로 MCF7 cell에 transfection 시킨 다음, HAT 배양액으로 선발하였으며, dexamethasone으로 유도시키고, 발현되는 정도를 면역 항체를 이용하여 분석하였다. 선발된 세포는 dexamethasone에 의하여 MMTV promoter가 유도되는 것을 확인할 수 있었다. 따라서 MCF7 cell과 같이 다양한 steroid receptor를 가지고 있는 세포는 유선 특정의 유전자 발현을 위한 세포 배양 모델에 유용하게 사용이 될 수 있을 것이다.