• Radiation Oncology Journal
• /
• 제19권4호
• /
• pp.389-396
• /
• 2001

목적 : 임상에서 사용하는 방사선량을 조사하여 환자의 자궁경부암 세포에서 유도되는 유전자를 검색하고자 하였다. 대상 및 방법 : 자궁경부암 환자에서 방사선치료 하루 전(대조군)과 1.8 Gy 조사후 40분 지나서(조사군) 자궁경부암 조직을 생검하여 각 군에서 total RNA를 추출하였다. differential display reverse transcription-polymerase chain reaction기법(DDRT-PCR)으로 발현이 증가 또는 감소된 유전자를 탐색하였다. 발현에 변화가 있는 cDNA를 추출하고 증폭하여 얻은 클론을 reverse Northern Blot방법을 이용하여 screening하였고, Northern Blot으로 확인하였다. sequencing을 실시한 후 NCBI database를 이용하여 blast search를 하였다. 발현이 감소한 유전자를 대상으로 다른 환자에서의 발현 양상을 확인하기 위하여 방사선치료를 받는 5명의 자궁경부암 환자를 대상으로 RT-PCR로 검사 하였다. 결과 : DDRT-PCR기법을 이용하여 방사선 조사군에서 발현이 증가 혹은 감소된 18개의 cDNA band를 발견하였다. reverse northern blot을 이용한 screening에서 발현이 증가된 10개의 클론과 감소된 1개의 클론을 확인하였다. 클로닝된 cDNA 조각들은 대부분 $400\~500\;bp$ 정도 되었으며, 그중 1개의 클론은 잘 알려져 있는 chemokine receptor CXCR4 유전자와 높은 상동성을 보였으며, 4개의 클론은 Human ESTs로 확인되었고 5개의 클론은 기능이 아직 확인되지 않은 알려져 있는 염기서열로 확인되었다. 방사선에 의하여 발현이 감소한 CxCa-11 클론이 모든 환자에서 방사선치료 전 시료에서 발현을 보였으나, 방사선치료 후 시료에서는 그 발현량이 감소하거나 발현이 안되었다. 결론 : DDRT-PCR을 이용하여 임상에서 자주 사용되는 방사선량을 환자의 자궁경부암에 조사했을 때 발현되는 유전자를 확인하였다. 이러한 유전자 발현의 확인은 방사선치료과정 중에 발생하는 일련의 기전들을 이해하는데 도움이 되리라 생각된다.

• BMB Reports
• /
• 제31권6호
• /
• pp.578-584
• /
• 1998

In a previous paper (Kim et al., 1996a), the immediate 5' -flanking region and coding region of the human UDP-N -acetylglucosamine:-D-mannoside-1,4-Nacetylglucosaminyltransferase III (N-acetylglucosaminyitransferase- III; GnT-III) gene was reported, isolated and analyzed. Herein, we report on amplification of a new 5' -noncoding region of the GnT-III mRNA by single-strand ligation to single-stranded cDNA-PCR (5' -RACE PCR) using poly(A)+ RNA isolated from human fetal liver cells. A cDNA clone was obtained with 5' sequences (96 bp) that diverged seven nucleotides upstream from the ATG (+1) start codon. A concensus splice junction sequence, TCTCCCGCAG, was found immediately 5' to the position where the sequences of the cDNA diverged. The result suggested the presence of an intron in the 5' -noncoding region and that the cDNA was an incompletely reversetranscribed cDNA product derived from an mRNA containing a new noncoding exon. When mRNA expression of GnT-III in various human tissues and cancer cell lines was examined, Northern blot analysis indicated high expression levels of GnT-III in human fetal kidney and brain tissues, as well as for a number of leukemia and lymphoma cancer cell lines. Promoter activities of the 5' -flanking regions of exon 1 and the new noncoding region were measured in a human hepatoma cell line, HepG2, by luciferase assays. The 5'-flanking region of exon 1 was the most active, whilst that of exon 2 was inactive.

• 생명과학회지
• /
• 제23권1호
• /
• pp.84-94
• /
• 2013

커피와 유사한 향미기를 갖는 녹차(Coffee-like green tea: CLGT)를 제조하고 이의 인체 유방암세포 주 MCF-7, 인체 전립선암세포 주 PC-3, 인체 신경모세포 주 SK-N-SH 및 쥐 심근세포 주 H9c2에 대한 세포독성을 연구하였다. 녹차엽(GTL)을 $240^{\circ}C$에서 1시간 roasting한 녹차가 커피와 가장 유사한 향미를 나타내었다. CLGT는 GTL과 비교하여 인체 암세포 및 정상세포와 쥐 심근세포에 대한 세포독성 차이가 없었다. GTL의 roasting은 ECG 등의 catechin 성분과 total protein은 유의성 있게 감소시켰지만, total phenol 및 total sugar는 유의성 있는 감소시키지 않아, 이와 같은 화합물 조성의 변화가 GTL에 커피유사 향미를 제공하였을 것이다. 따라서 이들 결과는 CLGT이 인체에 안전하여 커피 대용품으로 사용할 수 있음을 의미한다.

• Archives of Pharmacal Research
• /
• 제21권6호
• /
• pp.712-717
• /
• 1998

The complete nucleotide sequence of the cDNA clone encoding rat skeletal muscle myosin- binding protein H (MyBP-H) was determined and amino acid sequence was deduced from the nucleotide sequence (GenBank accession number AF077338). The full-length cDNA of 1782 base pairs(bp) contains a single open reading frame of 1454 bp encoding a rat MyBP-H protein of the predicted molecular mass 52.7kDa and includes the common consensus 1CA__TG' protein binding motif. The cDNA sequence of rat MyBP-H show 92%, 84% and 41% homology with those of mouse, human and chicken, respectively. The protein contains tandem internal motifs array (-FN III-Ig C2-FN III- Ig C2-) in the C-terminal region which resembles to the immunoglobulin superfamily C2 and fibronectin type III motifs. The amino acid sequence of the C-terminal Ig C2 was highly conserved among MyBPs family and other thick filament binding proteins, suggesting that the C-terminal Ig C2 might play an important role in its function. All proteins belonging to MyBP-H member contains `RKPS` sequence which is assumed to be cAMP- and cGMP-dependent protein kinase A phosphorylation site. Computer analysis of the primary sequence of rat MyBP-H predicted 11 protein kinase C (PKC)phosphorylation site, 7 casein kinase II (CK2) phosphorylation site and 4N-myristoylation site.

• Journal of Periodontal and Implant Science
• /
• 제29권3호
• /
• pp.663-676
• /
• 1999

Porphyromonas gingivalis, a Gram negative, anaerobic, asaccharolytic rod, is one of the most frequently implicated pathogens in human periodontal disease and has a requirement for hemin for growth. A 30 kDa (heated 24 kDa) hemin-binding protein whose expression is both hemin and iron regulated has recently been purified and characterized in this oral pathogen. This study has identified a hemin-binding P. gingivalis protein by expression of a P. gingivalis genomic library in Escherichia coli, a bacterium which does not require or transport exogenous hemin. A library of genomic DNA fragments from P. gingivalis was constructed in plasmid pUC18, transformed into Escherichia coli strain $DH5{\alpha}$ , and screened for recombinant clones with hemin-binding activity by plating onto hemin-containing agar. Of approximately 10,000 recombinant E. coli colonies screened on LB-amp-hemin agar, 10 exhibited a clearly pigmented phenotype. Each clone contained various insert DNA. The Hind III fragment transferred to the T7 RNA polymerase/promoter expression vector system produced a sligltly smaller (21 kDa) protein, a precursor form, immunoreactive to the antibody against the 24 kDa protein, suggesting that the cloned DNA fragment probably carried an entire gene for the 24 kDa hemin-binding protein.

• 한국발생생물학회:학술대회논문집
• /
• 한국발생생물학회 2003년도 제3회 국제심포지움 및 학술대회
• /
• pp.94-94
• /
• 2003

Rotavirus는 유아나 어린아이들에게 가장 일반적으로 나타나는 심한 위장염의 원인자이며 설사로 인한 심한 탈수 증세를 일으켜 급속히 성장하는 유아의 균형적인 영양 공급을 방해함으로써 유아들의 발육과 성장 그리고 심하면 생명에 커다란 영향을 미치게 된다. 한편 모유로 키운 유아들은 설사병의 낮은 발병율과 연관이 있었다. 특히 모유의 뮤신 복합체는 rotavirus에 특이적으로 결합하여 항 바이러스활동을 보여주는 것으로 나타났다. 이러한 배경에서 본 연구는 human breast tissue로부터 lactadherin의 cloning 및 sequence 분석을 통하여 유전자의 다양성을 조사하기로 하였다. 한국 여성 9명의 유두 근처 조직에서 lactadherin을 cloning하여 그 sequence를 보고 된 서양여성의 염기서열과 비교 분석결과 여러 곳에서 single nucleotide variation이 발견되었고 본 연구에서 클론한 lactadherin(31bp-1518bp)의 염기서열과 보고된 서양여성 lactadherin gene의 SNP와 비교하였을 때 8개의 SNP중 3부분만이 일치한다는 것을 확인하였다. 또한 같은 조직중 정상 조직과 암 조직 부분에서 각각 lactadherin을 클론하여 염기서열을 비교 분석하였는데 정상 조직에서 2곳의 silent mutation있었고 암조직에서 2곳의 mutation과 1곳의 silent mutation을 발견하였으며 전체 적으로 정상조직과 암 조직 부분에서 lactadherin을 clone하여 염기서열을 분석해본 결과 암조직일수록 유전자의 변이 비율이 높다는 것을 알 수 있었다. 그리고 동일한 염기서열 상에서 많은 변이가 일어났는데 286dp(A->C), 1418dp(G->C)은 mutation이었고 327dp (A->G), 454(C->T)은 silent mutation이었다. 그 외 DNA상에서 여러 부근에 변이가 존재하였는데 이 결과로 보아 coding region에 위치한 cSNP 중 amino acid 변화를 일으켜 protein structure 또는 function에 영향을 줄 수 있는 non-synonymous cSNP 일 것으로 예상되어지며 natural selection의 영향을 받고 있음을 암시하고 있다. 본 연구에서 관찰되어진 각각의 염기 서열의 변이는 한국 사람이 가지는 lactadherin gene의 cSNP의 일부라고 판단하였다.

• BMB Reports
• /
• 제29권2호
• /
• pp.146-150
• /
• 1996

A 2.1 kb cDNA clone for rat transketolase was isolated from rat liver ${\lambda}gt11$ cDNA library and its sequence was determined. The predicted rat transketolase (655 amino acids with $M_r$ 71,186) is highly similar (92%) to that of the human enzyme except that it contains an extra 32 amino acids at its N-terminus. Although it is less similar (<27%) to transketolases from non-mammalian species, the functional motifs such as the catalytic sites and thiamine binding domain are well conserved in the rat enzyme. Southern blot analysis of genomic DNA verified that transketolase appears to be derived from a single gene. Immunoblot and Northern blot analyses suggested that hepatic transketolase was activated pretranslationally by a 2.1-fold while little change was observed in brain enzyme, indicating a tissue-specific pretranslational activation during postnatal development.

• 한국의류학회지
• /
• 제24권8호
• /
• pp.1146-1154
• /
• 2000

The purpose of this study was to investigate the antimicrobial activity, antimutagenic and anticancer effects and dyeing properties of the fermented indigo extract. The methanol extract of fermented indigo showed a strong inhibition effect on Trich. mentagrophytes and antimutagenic activities against aflatoxin B$_1$(AFB$_1$) in the Ames test using Salmonella typhimurium TA 100. We also found in vitro anticancer effects of the methanol extract of fermented indigo and it was evaluated by using Clone M-3 mouse melanoma cells and A431 human epidermoid carcinoma cells and exerted little cytotoxity against 3T3-L1 embryo fibroblast cell. In the relationship between the K/S values of cotton and silk fabrics dyed with fermented indigo and dyeing repeating times, the K/S values became higher as the repeating times were increased. The K/S values were high when the fabrics were dyed at low temperture. The K/S values of cotton fabrics were higher than those of silk fabrics. Changes of surface color of silk fabrics were higher than those of cotton fabrics after water fastness test, laundering, irradiation and treatment of acidic and alkaline perspiration.

• Animal cells and systems
• /
• 제13권2호
• /
• pp.119-125
• /
• 2009

Mammalian hibernation is a type of natural adaptation that allows organisms to avoid harsh environment and to increase the possibility of survival. To investigate the molecular link between circadian and hibernating rhythms in the greater tube-nosed bats, Murina leucogaster, we set out to identify circadian genes that are expressed in bats, with specific focus on the PAS domain by using PCR-based screens. We could isolate a eDNA clone, designated as LPAS1, that encodes a protein of 521 amino acid residues. LPAS1 is closely related with CLOCK family with the highest homology to human CLOCK. Based on RT-PCR analyses, LPAS1 transcripts are ubiquitously present in tissues from both summer active and winter dormant periods. Given that LPAS1 is a member of the bHLH-PAS protein superfamily but lacks polyglutamine transactivation domains, it is likely to function as a repressor for endogenous CLOCK to hinder its roles in promoting transcription. Our result will open a new avenue to further examine the functional interconnection between the circadian clock and the circannual clock such as mammalian hibernation.

• 한국발생생물학회지:발생과생식
• /
• 제24권2호
• /
• pp.101-111
• /
• 2020

Coiled-coil domain containing 110 (CCDC110, KM-HN-1) is a protein containing C-terminal coiled-coil domain (CCD) which was previously discovered as a member of the human cancer/testis antigen (CTA). In addition, CCDC110 has both nuclear localization signal sequence and the leucine zipper motif. Although the functional role of CCDC110 has yet to be fully identified, the mRNA expression levels of CCDC110 are known to be highly elevated in various cancer types including testis, implying its relevance to cancer pathogenesis. In this study, we first developed several monoclonal antibody (mAb) hybridoma clones targeting CCDC110 and further isolated clone by characterizing for its specificity using immunoblotting and immunoprecipitation approaches with basal parenchymal sperm cells in testis tissue. Next, using these mAbs, we showed that the Tet-inducible overexpression of CCDC110 protein delayed the entry of G2/M phase in U2-OS osteosarcoma cells. Based on these results, we propose that CCDC110 plays a crucial role in cell cycle progression.