• Environmental Analysis Health and Toxicology
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• 제26권
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• pp.5.1-5.11
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• 2011

Objectives: In order to identify the possibility of slender bitterling (SB) (Acheilognathus yamatsutae) being used as a test species for estrogenic endocrine disrupting chemicals (EEDCs), we carried out the cloning and sequence characterization of the estrogen receptor (ER). Methods: The ER from a slender bitterling was obtained by reverse transcriptase-polymerase chain reaction (RT-PCR), 5'- and 3'-rapid amplification of cDNA ends (5'-RACE and 3'-RACE) and T-vector cloning. The expression of ER mRNA was also analyzed in six tissues (brain, liver, kidney, gill, gonad, and intestines) by real-time PCR. Results: We obtained an ER from the slender bitterling. The SB ER cDNA was 2189 base pairs (bp) in length and contained a 1707 bp open reading frame that encoded 568 amino acid residues. The SB ER amino acid sequence clustered in a monophyletic group with the $ER{\alpha}$ of other fish, and was more closely related to zebrafish $ER{\alpha}$(88% identity) than to the $ER{\alpha}$ of other fish. The SB ER cDNA was divided into A/B, C, D, E and F domains. The SB ER has conserved important sequences for ER functions, such as the DNA binding domain (D domain), which are consistent with those of other teleosts. Conclusions: The ER of the slender bitterling could provide basic information in toxicological studies of EEDCs in the slender bitterling.

• 한국임상수의학회지
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• 제32권4호
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• pp.352-355
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• 2015

본 연구는 암탐지에 우수한 능력을 보유하고 있는 공여견의 냄새 탐지 능력이 복제를 통하여 보존될 수 있을지를 알아보기 위하여 설계되었다. 직장암 탐지에 특화되어 훈련된 개를 복제하였고, 복제된 개는 환자와 건강한 지원자들로부터 채취된 호흡 샘플을 사용하여 유방암을 탐지하도록 훈련 되었다. 복제개의 암탐지 민감도는 93.3%, 특이도는 99.5%로 공여견의 암탐지 민감도 및 특이도 (91% 및 99%)와 유사하였다. 게다가 복제개는 유방암의 초기 단계까지 성공적으로 탐지할 수 있었다. 따라서 우수한 암탐지 능력은 복제를 통해서 보존될 수 있을 것이다.

• 한국동물학회:학술대회논문집
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• 한국동물학회 1999년도 한국생물과학협회 학술발표대회
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• pp.31-31
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• 1999

Characterization of mutant mice has been utilized as an animal model for the study of human inherited diseases. In addition to the pathogenesis stduy using the mutant mice, the mice have been used for the identification of the genes causing the phenotypes. Functional cloning and positional cloning are two approaches, depending on the phenotypes of the mutant mice. Though it takes a long time positional cloning has been well used to identify the gene of which function can not be presumed from the mouse phenotype. Recently by the advance of the molecular tools and the human genome project close to 10,000 genetic markers are developed to make the procedure faster. We obtained a new mutant mouse, sims, spontaneously arose and the affected mouse has a mild tremor and seizure was observed. Homozygote in either sex is sterile since uterus growth in female and seminal vesicle in male are not induced for the growth in puberty, implying the abnormal hormonal regulation during puberty. Supporting this, there is no detectable testosterone in the serum of the mutant male and the brain of the mutant is 30% heavier than littermate. To identify the location of the mutated gene, intraspecies cross to CAST/Ei was carried out and the 37 affected mice was analyzed for the linkage. The gene was mapped on chromosome 18, 20 cM from the centromere. More than 500 F2 progenies have been analyzed for the linkage and the locus becomes narrow within 3cM between Egrl and Fgf gene.f gene.

• 한국수산과학회지
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• 제42권2호
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• pp.139-145
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• 2009

We cloned and sequenced the open reading frame (ORF) cDNA encoding myostatin from the muscle of abalone (Haliotis discus hannai). The ORF cDNA of the abalone myostatin is 1134 bp and encoded 377 amino acid residues that were 60-96% homologous with the amino acids of other organism myostatins. In addition, the ORF contained a conserved proteolytic cleavage site (RXRR) and nine conserved cysteine residues in the C-terminus. Semi-quantitative RT-PCR revealed the presence of myostatin mRNA in various tissues. The strongest expression was observed in the mantle of female abalone, and the gills and heart of male abalone.

• Journal of Applied Biological Chemistry
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• 제51권3호
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• pp.95-101
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• 2008

The gene encoding a putative esterase of Xanthomonas oryzae pv. oryzae was cloned using PCR technique. The gene was expressed with His6 tag in E. coli. One-step purification of the recombinant esterase with Ni-NTA resin resulted in one band by SDS-PAGE analysis. The purified enzyme showed a molecular weight of 30 kDa, as expected, therefore the enzyme was a mononer. The enzyme was the most active toward p-nitrophenyl (p-NP) acetate and p-NP-butyrate to a lesser extent. However, the enzyme could not hydrolyze p-NP-myristate, palmitate, and stearate. Therefore, the enzyme is considered as an esterase, very different from lipase. The purified esterase had optimal pH at around 8.0 and was stable in a broad range of pH values. The optimal temperature ranged from 30 to $40^{\circ}C$, and the residual activity observed after heat treatment at $55^{\circ}C$ for 20 min was 72 % of the initial activity. The activity was inhibited by the presence of copper and cobalt ions.

• BMB Reports
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• 제39권3호
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• pp.311-318
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• 2006

One of the biggest group of proteins influenced by protein kinase CK2 is formed by factors engaged in gene expression. Here we have reported recently identified yeast transcription elongation factor Elf1 as a new substrate for both monomeric and tetrameric forms of CK2. Elf1 serves as a substrate for both the recombinant CK2$\alpha$' ($K_m$ 0.38 ${\mu}M$) and holoenzyme ($K_m$ $0.13\;{\mu}M$). By MALDI-MS we identified the two serine residues at positions 95 and 117 as the most probable in vitro phosphorylation sites. Co-immunoprecypitation experiments show that Elf1 interacts with catalytic ($\alpha$ and $\alpha$') as well as regulatory ($\beta$ and $\beta$') subunits of CK2. These data may help to elucidate the role of protein kinase CK2 and Elf1 in the regulation of transcription elongation.

• BMB Reports
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• 제40권3호
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• pp.444-447
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• 2007

Polymerase chain reaction (PCR) is a powerful technique in molecular biology and is widely used in various fields. By amplifying DNA fragments, PCR has facilitated gene cloning procedures, as well as molecular genotyping. However, the extraction of DNA from samples often acts as a limiting step of these reactions. In particular, the extraction of PCR-compatible genomic DNA from higher plants requires complicated processes and tedious work because plant cells have rigid cell walls and contain various endogenous PCR inhibitors, including polyphenolic compounds. We recently developed a novel solution, referred to as AnyDirect, which can amplify target DNA fragments directly from whole blood without the need for DNA extraction. Here, we developed a simple lysis system that could produce an appropriate template for direct PCR with AnyDirect PCR buffer, making possible the direct amplification of DNA fragments from plant leaves. Thus, our experimental procedure provides a simple, convenient, non-hazardous, inexpensive, and rapid process for the amplification of DNA from plant tissue.

• 미생물학회지
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• 제30권4호
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• pp.316-321
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• 1992

Catechol 2, 3-dioxygenase (C230) catalyses the oxidative ring cleavage of catechol to 2-hydroxymuconic semialdehyde. This is one of the key reactions in the metabolism of the widespresd pollutant aniline. We have cloned a gene encoding C230 from cells of the aniline degrading bacteria, Pseudomonas acidovorance KCTC2494 strain and expressed in E. coli, A 11.3-kilobase Sau3A partial digested DNA fragment from KCTC2494 was cloned into phagemid vector pBluescript and designated as pLP201. The C230 gene was mapped to a 2.8-kb region, and the derection of transcription was determined. The cloned C230 gene contains its own promoter which can be recognized and employed by E. coli transcriptional apparatus. C230 activities of subclones were identified by enzyme assay and activity staining. The T7 RNA promoter/polymerase system and maxicell analysis showed that a polypeptide with Mw of 35 kDa is the C230 gene product.

• Molecules and Cells
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• 제22권1호
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• pp.89-96
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• 2006

"Determinate" and "indeterminate" inflorescences in plants are controlled by a single recessive gene, for example, SELF-PRUNING (SP) in Solanum lycopersicum, TERMINAL FLOWER1 in Arabidopsis, CENTRORADIALIS in Antirrhinum, and CENTRORADIALIS-like gene in tobacco. Pepper (Capsicum annuum L.) is an indeterminate species in which shoots grow indefinitely. In this study, we cloned and characterized the pepper SP-like gene (CaSP). RT-PCR revealed that the CaSP transcript accumulates to higher levels in floral buds than in other organs. Comparison of genomic DNA and cDNA sequences from indeterminate and determinate pepper plants revealed the insertion of a single base in the first exon of CaSP in the determinate pepper plants. CaSP is annotated in linkage group 8 (chromosome 6) of the SNU2 pepper genetic map and showed similar synteny to SP in tomato. Transgenic tobacco plants overexpressing CaSP displayed late-flowering phenotypes similar to the phenotypes caused by overexpression of CaSP orthologs in other plants. Collectively, these results suggest that pepper CaSP is an ortholog of SP in tomato.

• 한국미생물·생명공학회지
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• 제45권3호
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• pp.226-235
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• 2017

Phytoene, lycopene, ${\beta}-carotene$과 같은 카로티노이드는 식품의 착색제나 영양보조제, 사료첨가제, 화장품의 원료로 사용된다. 이전 연구에서 본 연구진은 분홍색의 색소를 생산하는 새로운 해양 세균인 K. gwangalliensis를 분리 동정하였다. Phytoene desaturase (CrtI) 효소는 crtI 유전자에 암호화되어 있으며, phytoene을 lycopene으로 전환하며, 카로티노이드 합성 초기 단계에 있어서 필수적이다. CrtI는 카로티노이드 생합성 조절의 주요 효소 중 하나이며, 다양한 카로티노이드를 생합성하는 생물들의 카로티노이드 생합성 경로에 있어서 속도 조절 단계에 관련이 있다. 본 논문에서는 K. gwangalliensis로부터 lycopene 생합성을 담당하는 crtI 유전자를 클로닝 하였으며, 이 유전자는 1,584개의 염기서열을 가지며, 527개의 아미노산을 암호화하고 있다. crtI 유전자의 염기 서열을 Kocuria rhizophila와 Myxococcus xanthus를 포함한 다른 종의 염기 서열과 비교한 결과, 진화 과정에서 잘 보존되어 있음을 확인하였다. crtI 유전자를 포함하는 발현 플라스미드를 구축하여 발현시킨 결과, 이 플라스미드를 함유하는 대장균은 약 57 kDa의 재조합 단백질을 생산화였으며, 이는 phytoene desaturase의 분자량에 해당한다. lycopene의 생합성은 lycopene 생합성에 필요한 crtE, crtB 유전자를 포함한 pRScrtEB plasmid를 E. coli에 형질전환 했을 때, Escherichia coli에서 합성되는 것을 확인하였다. 이 연구의 결과는 분자 수준에서 K. gwangalliensis CrtI의 1차 구조에 대한 폭 넓은 지식 기반을 제공할 것이다.