• 한국수정란이식학회:학술대회논문집
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• 한국수정란이식학회 2002년도 국제심포지엄
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• pp.73-73
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• 2002

Nuclear transfer (NT) techniques have advanced in the last years, and cloned animals have been produced by using somatic cells in several species including pig. However, it is difficult that the nuclear transfer porcine embryos development to blastocyst stage overcoming the cell block in vitro. Abnormal segregation of chromosomes in nuclear transferred embryos on genome activation stage bring about embryo degeneration, abnormal blastocyst, delayed and low embryo development. Thus, we are evaluated that the correlations of the frequency of embryo developmental rates and chromosome aberration in NT and In viかo fertilization (IVF) derived embryo. We are used for ear-skin-fibroblast cell in NT. If only karyotyping of embryonic cells are chromosomally abnormal, they may difficultly remain undetected. Then, we evaluate the chromosome aberrations, fluorescent in situ hybridization (FISH) with porcine chromosome 1 submetacentric specific DNA probe were excuted. In normal diploid cell nucleus, two hybridization signal was detected. In contrast, abnormal cell figured one or three over signals. The developmental rates of NT and IVF embryos were 55% vs 63%, 32% vs 33% and 13% vs 17% in 2 cell, 8 cell and blastocyst, respectively. When looking at the types of chromosome aberration, the detection of aneuploidy at Day 3 on the embryo culture. The percentage of chromosome aneuploidy of NT and IVF at 4-cell stage 40.0%, 31.3%, respectively. This result indicate that chromosomal abnormalities are associated with low developmental rate in porcine NT embryo. It is also suggest that abnormal porcine embryos produced by NT associated with lower implantation rate, increase abortion rate and production of abnormal fetuses.

• BMB Reports
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• 제38권4호
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• pp.447-456
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• 2005

TNF-$\alpha$ plays a pivotal role in inflammation processes which are mainly regulated by endothelial cells. While TNF-$\alpha$ induces apoptosis of several cell types like tumor cells, endothelial cells are resistant to TNFa mediated cell death. The cytotoxic effects of TNF-$\alpha$ on most cells are only evident if RNA or protein synthesis is inhibited, suggesting that de novo RNA or protein synthesis protect cells from TNF-$\alpha$ cytotoxicity, presumably by NF-${\kappa}B$ mediated induction of protective genes. However, the cytoprotective genes involved in NF-${\kappa}B$ dependent endothelial cell survival have not been sufficiently identified. In the present study, the suppression subtractive hybridization (SSH) method was employed to identify rarely transcribed TNF-$\alpha$ inducible genes in human arterial endothelial cells related to cell survival and cell cycle. The TNF-$\alpha$-induced expression of the RNA binding protein $p54^{nrb}$ and the 14-3-3 protein HS1 as shown here for the first time may contribute to the TNF-$\alpha$ mediated cell protection of endothelial cells. These genes have been shown to play pivotal roles in cell survival and cell cycle control in different experimental settings. The concerted expression of these genes together with other genes related to cell protection and cell cycle like DnaJ, $p21^{cip1}$ and the ubiquitin activating enzyme E1 demonstrates the identification of new genes in the context of TNF-$\alpha$ induced gene expression patterns mediating the prosurvival effect of TNF-$\alpha$ in endothelial cells.

• 미생물학회지
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• 제30권4호
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• pp.286-290
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• 1992

광주근교 지역의 근권 토양으로부터 cetrimide agar medium을 이용하여 형광성 pseudomonads를 분리하였고, CAS medium에서 siderophore의 생성능력이 우수한 pseudomonads만을 분리하여 생리화학적인 실험을 수행하였다. Kanamycin-sensitive pseudomonads를 Tn5를 이용한 mutagenesis를 실시하여 Kanamycin에 내성을 갖는 transconjugants를 선별하였고, siderophore 생합성을 하지 못하는 돌연변이주를 선별하기 위하여 CAS medium에서 yellow hallow를 형성하지 못하거나 King's B medium에서 형광성을 나타내지 못하는 colony를 선별하였다. 선별된 mutants들의 genomic DNA에 Tn5가 삽입되었는지를 확인하기 위하여 Southern blot hybridization을 실시한 결과 intact Tn5에 homology를 나타내는 하나의 single band를 확인하였다.

• 한국미생물·생명공학회지
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• 제17권3호
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• pp.183-187
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• 1989

일반적으로 Mu phage를 가지고 있는 plasmid를 장내 세균에 삽입시키면 대부분의 Mu에 민감한 세균들은 zygotic induction이 일어나서 recipient cell 들이 살아남지 못하게 된다. 그러나 Mu 저항성 세균을 사용하면 cell이 죽지않고 recipient내에 삽입되는데 그 정확한 현상은 아직 밝혀지지 않았으나 Mu의 복제에 필요한 host의 기능이 결여된 것으로 추정되고 있다. 또한 reverse field electrophoresis를 사용하여 insertion mutant 나 deletion mutant들 의 염색체 및 거대 분자 DNA의 변이를 쉽게 비교 분석할 수가 있다. 본 실험에서는 Mu phage 저항성 C. crescentus를 사용하여 Tn5에 의한 영향 요구성 돌연변이주 출현률 및 운동성 돌연변이주 출현률을 조사한 결과 2%∼3% 수준으로 돌연변이가 일어났으며 이들 변이주들의 염색체를 Dra I 제한효소로 절단한 다음 reverse field electrophoresis로 분석한 결과 영양 요구성 돌연변이 균주들은 Tn5가 여러 위치에, 운동성에 돌연변이를 일으킨 균주들은 유사한 위치에 Tn5가 삽입된 것을 확인할 수 있었으나 hybridization 방법으로 확인한 것처럼 동시에 여러 위치를 확인할 수는 없었다. 그러나 이와 같은 문제들은 전기장의 교차시간 간격을 조절함으로 더 정확하게 확인할 수 있을 것으로 사료된다.

• Journal of Chest Surgery
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• 제44권2호
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• pp.123-130
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• 2011

Background: The aim of the present study was to identify chromosomal loci that contribute to the pathogenesis of aortic dissection (AD) in a Korean population using array comparative genomic hybridization (CGH) and to confirm the results using real-time polymerase chain reaction (PCR). Materials and Methods: Eighteen patients with ADs were enrolled in this study. Genomic DNA was extracted from individual blood samples, and array CGH analyses were performed. Four corresponding genes with obvious genomic changes were analyzed using real-time PCR in order to assess the level of genomic imbalance identified by array CGH. Results: Genomic gains were most frequently detected at 8q24.3 (56%), followed by regions 7q35, 11q12.2, and 15q25.2 (50%). Genomic losses were most frequently observed at 4q35.2 (56%). Real-time PCR confirmed the results of the array CGH studies of the COL6A2, DGCR14, PCSK6, and SDHA genes. Conclusion: This is the first study to identify candidate regions by array CGH in patients with ADs. The identification of genes that may predispose an individual to AD may lead to a better understanding of the mechanism of AD formation. Further multicenter studies comparing cohorts of patients of different ethnicities are warranted.

• 한국수정란이식학회지
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• 제24권2호
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• pp.77-87
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• 2009

To understand molecular and cellular mechanisms of many gene products in the female reproductive organs including the ovary and uterine endometrium as well as during embryo development, researchers have developed and utilized many effective methodologies to analyze gene expression in cells, tissues and animals over the last several decades. For example, blotting techniques have helped to understand molecular functions at DNA, RNA and protein levels, and the reverse transcription-polymerase chain reaction (RT-PCR) method has been widely used in gene expression analysis. However, some conventional methods are not sufficient to understand regulation and function of genes expressed in very complex patterns in many organs. Thus, it is required to adopt more high-throughput and reliable techniques. Here, we describe several techniques used widely recently to analyze gene expression, including annealing control based-PCR, differential display-PCR, expressed sequence tag, suppression subtractive hybridization and microarray techniques. Use of these techniques will help to analyze expression pattern of many genes from small scale to large scale and to compare expression patterns of genes in one sample to another. In this review, we described principles of these methodologies and summarized examples of comparative analysis of gene expression in female reproductive organs with help of those methodologies.

• 한국응용곤충학회지
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• 제38권2호
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• pp.145-149
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• 1999

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.

• 대한기관식도과학회지
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• 제5권1호
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• pp.49-54
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• 1999

The etiology of inverted papilloma(IP) remains unknown, but several studies have reported that Human Papillomavirus(HPV) may play a role in the pathogensis of sinonasal inverted papilloma(IP). And recent reports demonstrate the possible etiologic role of Epstein-Barr virus (EBV) in sinonasal IP. The aim of this study is to detect HPV and EBV in sinonasal IP, to examine the relationship between HPV subtype and sinonasal IP, to investigate the relation between HPV and EBV. We reviewed 30 cases of sinonasal IP(simple IP 19 cases, IP with dysplasia 8 cases, IP with squamous cell carcinoma 3 cases). Paraffin embedded archival tissue was used in this study. Detection of HPV, EBV were examined by in situ hybridization(ISH) using HPV type 6/11, 16/18, 31/33/35 DNA probe and EBER probe. The HPV was detected in 6(20%) out of 30 cases. The HPV 6/11 was dectected in 4 out of 19 cases of simple IP, HPV 16/18 in 1, HPV 31/33/35 in 1 out of 8 cases of IP with dysplasia respectively. The EBV was not detected in 30 cases. HPV may play a role in the pathogensis of sinonasal inverted papilloma. But EBV is not a etiopathologic factor to be considered in the development of sinonasal IP.

• 미생물학회지
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• 제26권4호
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• pp.305-311
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• 1988

Slow growing R. japonicum R-l68 균주로부터 spectinomycin 내성 균주를 선발하고 이 Rhizobium내에 Tn-5를 도입시키기 위하여 Tn5가 함유된 E. coli WA 803/pGS9과의 conjugation을 통한 transposon mutagenesis를 실시하였다. 이때 C conjugation을 통한 Tn5 전이 빈도는 $1.0\times 10^{-5}-5.0\times 10^{-7}$ 범위 이였으며, 얻어진 transconjugant들은 spectinomycin (($100{\mu}$g/ml)과 kanamycin ($50{\mu}$g/ml)을 함유한 yeast extract-mannitol 배지에서 8-10일 배양후 colony를 형성하였다. 또한 transconjugant들은 genome상에 Tn-5를 함유하고 있음을 hybridization-을 통하여 확인하였다. 한편 nodule은 형성 하나 질소고정 활성이 없는 돌연변이주 R. japonicum RMa 75 $nod^{+}fix^{-}$ 균주를 선발하였는데 이 균주는 nodule내에 leghemoglobin이 결핍되어 있음이 확인되었다.

• 미생물학회지
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• 제40권1호
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• pp.60-64
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• 2004

Gentamicin에 저항성을 나타내는 세균을 병원 하수로부터 분리하여 aminoglycoside-(3)- N-acetyltransferase를 암호화하는 유전자인 aac(3)II의 존재여부를 dot-blot hybridization으로 조사하였다. aac(3)II유전자의 일부를 탐침으로 사용한 결과 gentamicin저항성 세균의 41% (39/95)가 이 유전자를 가지고 있었다. aac(3)II와 Tn3에서 각각 설계된 primer를 사용한 PCR 결과 aac(3)II를 가지고 있는 39개 균주 중 13개 균주가 TnS-aac(3)II구조를 가지고 있음을 알 수 있었다. aac(3)II의 상류에 Tn3를 가지고 있는 13개 균주의 gentamicin에 대한 최소억제농도는 가지고 있지 않은 균주에 비해 상대적으로 높았다. 13개 균주를 동정한 결과 5개는 Eschelichia coli, 3개는 Acinetobacter johnsonii, Enterobacter agglomerans와 Micrococcus luteus가 각각 2개, 그리고 1개의 Pseudomonas facilis로 동정되었다. 이러한 결과들은 Tn3-aac(3)II구조가 gentamicin 저항성 세균들에서 널리 분포하고 있음을 말해준다.