• Journal of Plant Biotechnology
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• 제45권4호
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• pp.322-327
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• 2018

Post-translational modification of proteins regulates signaling cascades in eukaryotic system, including plants. Among these modifications, phosphorylation plays an important role in modulating the functional properties of proteins. Plants perceive environmental cues that directly affect the phosphorylation status of many target proteins. To determine the effect of environmentally induced phosphorylation in plants, in vivo methods must be developed. Various in vitro methods are available but, unlike in animals, there is no optimized methodology for detecting protein phosphorylation in planta. Therefore, in this study, a robust, and easy to handle Phos-Tag Mobility Shift Assay (PTMSA) is developed for the in vivo detection of protein phosphorylation in plants by empirical optimization of methods previously developed for animals. Initially, the detection of the phosphorylation status of target proteins using protocols directly adapted from animals failed. Therefore, we optimized the steps in the protocol, from protein migration to the transfer of proteins to PVDF membrane. Supplementing the electrophoresis running buffer with 5mM $NaHSO_3$ solved most of the problems in protein migration and transfer. The optimization of a fast and robust protocol that efficiently detects the phosphorylation status of plant proteins was successful. This protocol will be a valuable tool for plant scientists interested in the study of protein phosphorylation.

• 한국환경성돌연변이발암원학회지
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• 제23권2호
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• pp.56-62
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• 2003

The liver progenitor cells could form a potential target cell population fore both tumor-initiating and -promoting chemicals. Induction of drug-metabolizing and antioxidant enzymes, including AhR-dependent CYP1A1, NQO-1 and AKR1C9, was detected in the rat liver epithelial WB-F344 "stem-like" cells. Additionally, WB-F344 cells express a functional, wild-type form of p53 protein, a biomarker of genotoxic events, and connexin 43, a basic structural unit of gap junctions forming an important type of intercellular communication. In this cellular model, two complementary assays have been established for detection of the modes of action associated with tumor promotion: inhibition of gap junctional intercellular communication (GJIC) and proliferative activity in confluent cells. We found that the PAHs and PCBs, which are AhR agonists, released WB-F344 cells from contact inhibition, increasing both DNA synthesis and cell numbers. Genotoxic effects of some PAHs that lead to apoptosis and cell cycle delay might interfere with the proliferative activity of PAHs. Contrary to that, the nongenotoxic low-molecular-weight PAHs and non-dioxin-like PCB congeners, abundant in the environment, did not significantly affect cell cycle and cell proliferation; however both groups of compounds inhibited GJIC in WB-F344 cells. The release from contact inhibiton by a mechanism that possibly involves the AhR activation, inhibition of GJIC and genotoxic events induced by environmental contaminants are three important modes of action that could play an important role in carcinogenic effects of toxic compounds. The relative potencies to inhibit GJIC, to induce AhR-mediated activity, and to release cells from contact inhibition were determined for a large series of PAHs and PCBs and their metabolites. In vitro bioassays based on detection of events on cellular level (deregulation of GJIC and/or proliferation) or determination of receptor-mediated activities in both ?$stem-like^{\circ}{\times}$ and hepatocyte-like liver cellular models are valuable tools for detection of modes of action of polyaromatic hydrocarbons. They may serve, together with concentration data, as a first step in their risk assessment.

• 대한소아치과학회지
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• 제33권2호
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• pp.207-220
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• 2006

새로 개발된 조기 진단 장비들로는 laser fluorescence device(LFD), 초음파 진단 시스템, confocal laser scanning microscopy(CLSM), quantitative light-induced fluorescence(QLF) 시스템 그리고 digital imaging fiber-optic trans-illumination(DIFOTI) 시스템 등이 있다. 본 연구는 임상에서 사용되고 있는 DIFOTI시스템과 LFD를 이용하여 유치 교환 시기에 있는 환아 21명, 25개의 유치를 대상으로 각 치아당 $1{\sim}3$점을 선정하여 구강내에서 초기 우식법랑질에 대한 DIFOTI 이미지 촬영 결과와 LFD 계측값을 3회 측정하고 그 평균을 대표값으로 한 결과를 CLSM과 비교하여 진단 능력을 평가하였다. 실험실 연구에서는 인공우식 용액을 이용하여 수거된 40개의 유치를 협설면에 $2{\times}3mm$ 크기의 창을 형성하고 4, 8, 12, 16일간 탈회시키면서 그 변화를 DIFOTI 시스템과 LFD를 이용하여 측정하고 이를 CLSM과 비교 분석하여 다음과 같은 결론을 얻었다. 1. 구강내에서 촬영된 DIFOTI의 민감도는 88.2%이었고, 특이도는 76.9%이었다. 2. 구강내에서 측정된 LDD의 민감도는 76.5%이었고, 특이도는 69.2%이었다. 3. 실험실 연구에서 유치 법랑질의 탈회 기간에 따른 DIFOTI 광투과율의 회귀 분석한 결과, 탈회 시간에 따라 광투과율은 감소하였다(r=-0.96, p<0.05). 4. 실험실에서 탈회 기간에 따른 LFD의 측정값의 회귀 분석 결과 통계적 유의성을 보이지 않았다(p>0.05). 5. DIFOTI 이미지의 광투과율과 CLMS의 병소 깊이에 대한 상관 계수는 -0.688이었으나(p<0.05), LFD의 측정값은 유의성을 보이지 않았다.

• Pediatric Infection and Vaccine
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• 제3권2호
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• pp.133-138
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• 1996

Purpose : In the treatment of MRSA infection, rapid detection of MRSA is extremely important. The mecA gene codes the new drug resistant polypeptides called PBP2' which mediates the clinically relevant resistance to all beta-lactam antibiotics. The identical mecA gene has been found in coagulase-negative staphylococcus with the methicillin-resistant phenotype. On the other hand, the femA gene was absent from coagulase negative staphylococcus strains with the methicillin resistant phenotype. This study is aimed at early detection and definite diagnosis of MRSA. Methods : A total of 24 MRSA strains were studied. All strains were tested for antimicrobial susceptibility and purified DNA. We amplified both mecA and femA genes by PCR in 24 strains. Results : In MRSA all the 16 strains (100%) carried femA gene and 11 strains (68.7%) carried mecA gene. In contrast, in methicillin sensitive staphylococcus all the 8 strains (100%) carried femA and only 3 strains (37.5%) were detected mecA. Conclusions : As results, there are difference in the phenotype and genotype of methicillin resistance by PCR of mecA and femA. Such disparities between methicillin resistance and the presence of mecA gene suggest the presence of control gene of the mecA.

• The Plant Pathology Journal
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• 제32권1호
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• pp.53-57
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• 2016

Major diseases in grafted cacti have been reported and Fusarium oxysporum, Bipolaris cactivora, Phytophthora spp. and Collectotrichum spp. are known as causal pathogens. These pathogens can lead to plant death after infection. Therefore, some European countries have quarantined imported cacti that are infected with specific fungal pathogens. Consequently, we developed PCR detection methods to identify four quarantined fungal pathogens and reduce export rejection rates of Korean grafted cacti. The pathogen specific primer sets F.oF-F.oR, B.CF-B.CR, P.nF-P.nR, and P.cF-P.CR were tested for F. oxysporum, B.cactivora, P. nicotinae, and P. cactorum, respectively. The F.oF-F.oR primer set was designed from the Fusarium ITS region; the B.CF-B.CR and P.nF-P.nR primers respectively from Bipolaris and Phytophthora ITS1; and the P.cF-P.CR primer set from the Ypt1protein gene region. The quarantine fungal pathogen primer pairs were amplified to the specific number of base pairs in each of the following fungal pathogens: 210-bp (F. oxysporum), 510-bp (B. cactivora), 313-bp (P. nicotinae), and 447-bp (P. cactorum). The detection limit for the mono- and multiplex PCR primer sets was 0.1 ng of template DNA under in vitro conditions. Therefore, each primer set successfully diagnosed contamination of quarantine pathogens in export grafted cacti. Consequently, our methodology is a viable tool to screen contamination of the fungal pathogen in exported grafted cacti.

• 대한화장품학회지
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• 제42권3호
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• pp.211-216
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• 2016

소핵시험은 세포분열 단계 중 간기 세포의 세포질 내 소핵 유무를 조사함으로써 유전독성을 평가하는 시험법이다. 최근 화장품 안전성 평가에 동물실험을 금지하거나 최소화하려는 노력이 확산되고 있어 유전독성 평가에 있어서도 기존의 동물실험이 아닌 새로운 in vitro 시험법이 요구되고 있다. 본 연구에서는 3차원 배양인공피부모델인 KeraSkin$^{TM}$을 이용하여 도포 처치된 물질의 유전독성을 평가하였다. 2종의 유전독성물질인 mitomycin C (MMC)와 methyl methanesulfonate (MMS)는 농도 의존적으로 세포독성과 소핵 형성이 유도된 반면, 대조물질인 4-nitrophenol (4-NP)와 trichloroethylene (TCE)에서는 농도 의존적으로 세포독성은 관찰되었으나 소핵은 형성되지 않았다. 따라서 인공피부모델을 이용한 소핵시험이 화장품과 같은 피부적용물질의 in vitro 유전독성 평가에 유용할 것으로 사료된다.

• 한국환경성돌연변이발암원학회지
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• 제19권1호
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• pp.14-19
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• 1999

Chromosome rearrangements induced in human lymphocyte after in vitro exposure to chromium were analysed by the use of fluorescence in situ hybridization(FISH) with triple combination of composite whole chromosome-specific probe for chromosome 1, 2 and 4. Chromosome aberrations was scored by the Protocol for Aberration Identification and Nomenclature Terminology (PAINT). Stable translocation was the most frequent type of aberrations and dicentrics and insertions were also observed. Chromium treatment enhanced the frequencies of stable translocations and color junctions in a dose-dependent manners, but no distinct increase of dicentrics and insertions was seen. The ratio of the yields of translocation to the yields of dicentric varied between 13 to 27. The presents results demonstrate fluorescent in situ hybridization (FISH) is useful for detecting chromosomal rearrangements induced by chromium.

• 센서학회지
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• 제32권6호
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• pp.403-411
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• 2023

The shift in the medical paradigm from treatment to prevention and diagnosis has underscored the growing significance of biosensors. Notably, the recent COVID-19 pandemic has spurred the widespread adoption of biosensors for the detection of viral genes and antigens. Consequently, there has been a substantial increase in both the demand for biosensors and the industries associated with their production. Furthermore, biosensors find applications not only in healthcare but also in diverse fields such as environmental monitoring, food quality control, military defense, and industrial processes. In this brief review, we delve into the essential attributes of biosensors, namely sensitivity, selectivity, and stability. We provide an overview of the latest research trends aimed at improving these attributes. Additionally, we introduce recent research cases in which these attributes are being applied both in vivo and in vitro.

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2005년도 생물공학의 동향(XVI)
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• pp.17-17
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• 2005

In this presentation, we will discuss several recent achievements developed in my laboratory for microarray-based diagnosis of human genetic mutations including HNF-1 and BRCA1 mutations. To determine the presence of the genetic mutations in a human sample, we prepared allele-specific oligonucleotide chips from selected mutation sites and generated target probes using a tow-step method for Cy-3 DNA $samples^{1)}$ or in vitro transcription of promoter-tagged PCR products for Cy-3 RNA $samples^{2)}$. Hybridization of the target probes to the chips successfully identified all of the genotypes for the tested sites. For more reliable diagnosis, we also employed single base extension (SBE) reaction and zip-code microarray technique for our strategy. Particularly we developed an efficient PNA zip-code microarray for the detection of $HNF-1{\alpha}$ $mutations^{3)}$. Using multiplex SBE reactions and zip-code strategy, we were able to correctly diagnose several mutation sites in exon 2 of $HNF-1{\alpha}$ with a wild-type and mutant including a MODY3 patient. These works represent successful applications of DNA microarray technology for the diagnosis of human genetic mutations.

• Applied Science and Convergence Technology
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• 제23권2호
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• pp.61-71
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• 2014

A field-effect transistor (FET) is one of the most commonly used semiconductor devices. Recently, increasing interest has been given to FET-based biosensors owing totheir outstanding benefits, which are likely to include a greater signal-to-noise ratio (SNR), fast measurement capabilities, and compact or portable instrumentation. Thus far, a number of FET-based biosensors have been developed to study biomolecular interactions, which are the key drivers of biological responses in in vitro or in vivo systems. In this review, the detection principles and characteristics of FET devices are described. In addition, biological applications of FET-type biosensors and the Debye length limitation are discussed.