북방종개(Iksookimia pacifica)의 유전적 다양성과 구조적 특징을 밝히기 위해 동해 독립하천들에 서식하는 10개의 집단들을 대상으로 핵유전자와 미토콘드리아 유전자에 기반한 집단유전학적 분석을 실시하였다. 일부 예외적인 경우를 제외하고, 미토콘드리아와 핵유전자 모두 통계적으로 유의미한 집단 간 유전적 분화가 관찰되었다. 핵유전자들의 DNA 서열자료에서 추출한 유전자형 자료를 Bayesian 방법으로 분석한 결과 북방종개는 천진천과 양양남대천을 기준으로 북쪽과 남쪽의 두 개의 그룹으로 나뉘는 구조를 보였다. 현재 동해 하천들이 지리적으로 단절되어 온 독립 수계라는 것을 감안했을 때, 남북으로 구별되는 집단유전적 구조는 북방종개가 한반도에 정착했던 초기 조상 집단이 남북으로 갈라지는 지리적인 분리 사건과 관련되었을 것으로 해석되며, 이러한 초기 조상집단의 지리적 분리 이후, 두 조상 그룹들은 남북의 지리적인 범위 내에서 하천 별 고립에 따른 추가적인 분화 과정을 거쳤을 것으로 추정된다. 주목할 점으론, 자산천 집단의 많은 개체들이 지리적 거리가 먼 양양남대천 및 강릉남대천 집단과 하나의 유전적 cluster를 형성하고 있는 것이다. 이와 함께 미토콘드리아 유전자의 경우 몇몇 이웃하는 집단들 사이에 현저히 낮은 유전적 분화도 그리고 일부 집단들에서 매우 낮은 유전적 다양성이 관찰되었다. 본 집단유전학적 결과는 향후 북방종개의 보존 및 관리를 위한 기초자료로 제시될 것이다.

The purpose of this study is to develop transgenic cell line expressing targeted human granulocyte colony stimulating factor (hGCSF) and green fluorescence protein (GFP) genes as well as production of Somatic Cell Nuclear Transfer (SCNT) embryos derived from co-expressed transgenic donor cells. Constructed pPiggy-mWAP-hGCSF-EF1-GFP vector was chemically transfected into bovine fetus cells and then, only GFP expressed cells were selected as donor cells for SCNT. Cleavage and blastocyst rates of parthenogenetic, SCNT embryos using non-TG cell and hGCSF-GFP dual expressed SCNT embryos were examined (cleavage rate: 78.0±2.8 vs. 73.1±3.2 vs. 70.4±4.3%, developmental rate: 27.2 ±3.2 vs. 21.9±3.1 vs. 17.0±2.9%). Result indicated that cleavage and blastocyst rates of TG embryos were significantly lower (P<0.05) than those of parthenogenetic and non-TG embryos, respectively. In this study, we successfully produced hGCSF-GFP dual expressed SCNT embryos and cryopreserved to produce transgenic cattle for bioreactor system purpose. Further process of our research will transfer of transgenic embryos to recipients and production of hGCSF secreting cattle.

This study aimed at investigating the gene expression profile in basal ganglia of hexa-valence chromium exposed rat based on cDNA array analysis. For cDNA array, Sprague-Dawley male rats (300 ± 25 g) were administrated with 15 mg/kg B.W/day of potassium dichromate by gavage (0.3 ml) dissolved in saline for 10 days (n=5). For dose-related gene expression analysis, rats were administrated with 0, 1, 5 mg/kg B.W/day of potassium dichromate for 10 days. Control rats were administrated with equal volume of saline (n=5). For cDNA array analysis, RNA samples were extracted from brain tissue and reverse-transcribed in the presence of [α³²P]-dATP. Membrane sets of the Atlas array II and Toxicology array kits were hybridized with cDNA probe sets. RT-PCR and Northern blot hybridization methods were employed for validation and assessment of the dose-related gene expression profile, respectively. Among the 2352 cDNAs, 43 genes showed significant (>two-fold) changes in expression. 22 genes were up-regulated and 21 genes were down-regulated in the 15 mg/kg B.W/day hexa-valence chromium treated group than control. According to the Northern blot hybridization analysis, heat shock protein 47, neurodegeneration associated protein 1 and pituitary specific growth factor 1a genes were up-regulated, but Gamma-aminobutyl-acid a1 subunit, neuroligin2, brain calcium-transporting plasma membrane type ATPase genes were down-regulated even in the low-dose of hexa-valence chromium exposed group (1 mg/kg B.W/day) than control. Genes that detected in this study may be closely related to the hexa-valence chromium-induced neurotoxicity in the rat basal ganglia and addition study of these genes can give some more useful information about the neuro-toxic mechanism by hexa-valence chromium.

야생주 핵다각체병 바이러스의 낮은 병원성에 대해 살충제로서의 파밤나방 핵다각체병 바이러스(Spodoptera exigua nucleopolyhedrovirus: SeNPV)의 병원성 향상을 꾀함과 동시에 재조합 바이러스가 다각체 내에 매립됨으로써 살충제로의 적용시 야외에서의 안정성을 확보할 수 있는 p10 유전자의 프로모터를 이용한 새로운 발현벡터를 개발하기 위하여 국내분리주 SeNPV의 p10 유전자 구조를 분석하였다. 그 결과, SeNPV p10 유전자의 프로모터와 구조유전자 부위를 포함한 545염기서열을 결정하여 기존에 보고된 SeNPV p10 유전자(Zuidema et al., 1993)와 비교한 결과 구조유전자 부위에서는 100%의 상동성을 보였으나 5` 및 3` flanking region의 4개의 염기서열에서 차이를 보였다. Southern hybridization에 의하여 SeNPV전체 genomic DNA상에서 p10 유전자는 Sph I 2.4Kb와 Cla I 4.0Kb 단편내에 각각 존재함을 확인하였으며, p10 유전자를 포함하는 이들 단편을 각각 클로닝하여 제한효소 지도를 작성한다.

The pronuclear injection of metallothionein-human growth hormone (MT-hGH) gene into rabbit zygotes was performed to establish in vitro developmental system and to detect the presence of the injected gene by nested PCR. Mature female New Zealand White rabbits were superovulated by eGG and hCG treatments. The rabbits were mated and the zygotes were collected from the oviducts 18-22 h after hCG injection by flushing with D-PBS. Two to three picoliters of MT-hGH gene was microinjected into male pronuclei. The foreign gene-injected zygotes were cultured in TCM-199 or RD mediurn containing 10% FCS with a monolayer of rabbit oviductal epithelial cefls in a 5% incubator. The presence of injected DNA in rabbit embryos or blastomeres at different developmental stages .vas detected by a nested PCR analysis. The results are summarized as follows ; 1.The developmental rate of the MT-hGH gene-injected zygotes to blastocyst was significantly higher in TCM-199 medium (68.1%) than in RD medium (42.9%). 2.The gene injection into pronuclei at 18 or 22 hours post hCG treatment during pronuclear stage did not much affect on the in vitro development of the rabbit embryos. 3.The rate of gene-positive embryos detected by the nested PCR analysis was significantly decreased when they developed to blastocysts. The results indicate that the screening of transgene in rabbit embryos by nested PCR analysis could be a prornisible method for the preselection of transgenic embryos. Furthermore, the preselection of transgenic embryos would greatly reduce hoth the cost and effort of production of transgenic animals.

흰 불나방 핵 다각체바이러스(Hyphantria cunea nuclear Palyhedrosis virus: HcNPV) 다각체 단백질 유전자포함 절편의 탐색과 클로닝을 행하였다. Autographa cahfornica NPV EeoRI-I 절편 (약 7.3 kb), Bombyx mori NPV PstI-F 절편 (약 7 kb) 및 합성 oligonucleotide ( 3D-mer) 를 probe로 한 southern hybridization을 행하여 HcNPV PstI - L 절편 (5.3 kb)을 탐색하고, pUC18을 이용하여 E. coli에 형질전환시켜 클로닝하였다. 클로닝한 plasmid의 EeoRI, SaIl, Kpnl, HindIII, SacI 및 AvaI의 제한효소지도를 작성하고 pHeP-L(8.0 kb)이라 명명히였으며, 이를 다시 pHcP-Ll(4.7 kb), pHcP-L2(7.1 kb), pHcP-L3(5.3 kb), pHcP-L4(4.2 kb) 및 pHeP-L5(4.5 kb)로 subeloning 하였다.