• 제목/요약/키워드: Molecular Detection

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Use of DNA Methylation for Cancer Detection and Molecular Classification

  • Zhu, Jingde;Yao, Xuebiao
    • BMB Reports
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    • 제40권2호
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    • pp.135-141
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    • 2007
  • Conjugation of the methyl group at the fifth carbon of cytosines within the palindromic dinucleotide 5'-CpG-3' sequence (DNA methylation) is the best studied epigenetic mechanism, which acts together with other epigenetic entities: histone modification, chromatin remodeling and microRNAs to shape the chromatin structure of DNA according to its functional state. The cancer genome is frequently characterized by hypermethylation of specific genes concurrently with an overall decrease in the level of 5-methyl cytosine, the pathological implication of which to the cancerous state has been well established. While the latest genome-wide technologies have been applied to classify and interpret the epigenetic layer of gene regulation in the physiological and disease states, the epigenetic testing has also been seriously explored in clinical practice for early detection, refining tumor staging and predicting disease recurrence. This critique reviews the latest research findings on the use of DNA methylation in cancer diagnosis, prognosis and staging/classification.

Development of High-Field ESR System Using SQUID Magnetometer and its Application to Measurement under High Pressure

  • Sakurai, T.;Fujimoto, K.;Okubo, S.;Ohta, H.;Uwatoko, Y.
    • Journal of Magnetics
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    • 제18권2호
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    • pp.168-172
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    • 2013
  • We have developed a high-field and high-frequency ESR system using a commercially available magnetometer equipped with the superconducting quantum interference device (SQUID). This is magnetization detection type ESR and ESR is observed as a change of the magnetization at the resonance condition under irradiation of the electromagnetic wave. The frequency range is from 70 to 315 GHz and the maximum magnetic field is 5 T. The sensitivity is estimated to be $10^{13}$ spins/G. The advantage of this system is that the high-field ESR measurements can be made very easily and quantitatively. Moreover, this high-field ESR can be applied to the measurements under pressure by using a widely used piston-cylinder pressure cell.

MEASUREMENT OF PESTICIDES RESIDUES USING SPECTROSCOPY ON AGRICULTURAL PRODUCTS

  • Kim, Y. W.;S. H. Noh
    • 한국농업기계학회:학술대회논문집
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    • 한국농업기계학회 2000년도 THE THIRD INTERNATIONAL CONFERENCE ON AGRICULTURAL MACHINERY ENGINEERING. V.III
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    • pp.525-532
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    • 2000
  • A new spectroscopic method for pesticide residues detection on agricultural products was developed. The general determination methods are high performance liquid chromatography (HPLC), gas chromatography (GC) or GC-mass spectrometry. They have provided relatively good detection limit and accuracy with complicated and time-consuming (5hrs above) procedures. In addition freshness is very important for evaluating qualities of agricultural products. This requires a simple and fast method for detection of pesticides. Reflectance, transmittance and fluorescence spectrometry of pesticides were tested using UV range because most of pesticides contain conjugation band in the molecular structures. Fluorescence spectrometry showed better sensitive to detect pesticide residues than did reflectance and transmittance spectrometry. Intensity and shape of fluorescence spectra showed different patterns with different structures of pesticides. Detection limit for fluorescence spectrometry was 0.1 ppm to 10 ppm depending on the structures of pesticides. Application of fluorescence spectrometry appears to be an easy method for detection of pesticide residues on agricultural products.

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Noninvasive fetal RHD genotyping using cell-free fetal DNA incorporating fetal RASSF1A marker in RhD-negative pregnant women in Korea

  • Han, Sung-Hee;Yang, Young-Ho;Ryu, Jae-Song;Kim, Young-Jin;Lee, Kyoung-Ryul
    • Journal of Genetic Medicine
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    • 제12권2호
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    • pp.100-108
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    • 2015
  • Purpose: Conventional methods for the prenatal detection of fetal RhD status involve invasive procedures such as fetal blood sampling and amniocentesis. The identification of cell-free fetal DNA (cffDNA) in maternal plasma creates the possibility of determining fetal RhD status by analyzing maternal plasma DNA. However, some technical problems still exist, especially the lack of a positive control marker for the presence of fetal DNA. Therefore, we assessed the feasibility and accuracy of fetal RHD genotyping incorporating the RASSF1A epigenetic fetal DNA marker from cffDNA in the maternal plasma of RhD-negative pregnant women in Korea. Materials and Methods: We analyzed maternal plasma from 41 pregnant women identified as RhD-negative by serological testing. Multiplex real-time PCR was performed by amplifying RHD exons 5 and 7 and the SRY gene, with RASSF1A being used as a gender-independent fetal epigenetic marker. The results were compared with those obtained by postnatal serological analysis of cord blood and gender identification. Results: Among the 41 fetuses, 37 were RhD-positive and 4 were RhD-negative according to the serological analysis of cord blood. There was 100% concordance between fetal RHD genotyping and serological cord blood results. Detection of the RASSF1A gene verified the presence of cffDNA, and the fetal SRY status was correctly detected in all 41 cases. Conclusion: Noninvasive fetal RHD genotyping with cffDNA incorporating RASSF1A is a feasible, reliable, and accurate method of determining fetal RhD status. It is an alternative to amniocentesis for the management of RhD-negative women and reduces the need for unnecessary RhIG prophylaxis.

A Rapid Preconcentration Method Using Modified GP-MSE for Sensitive Determination of Trace Semivolatile Organic Pollutants in the Gas Phase of Ambient Air

  • He, Miao;Xu, Qingjuan;Yang, Cui;Piao, Xiangfan;Kannan, Narayanan;Li, Donghao
    • Bulletin of the Korean Chemical Society
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    • 제35권10호
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    • pp.2995-3000
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    • 2014
  • A sensitive concentration method utilising modified gas-purge microsyringe extraction (GP-MSE) was developed. Concentration (reduction in volume) to a microlitre volume was achieved. PAHs were utilised as semivolatile analytes to optimise the various parameters that affect the concentration efficiency. The injection rate and temperature were the key factors that affected the concentration efficiency. An efficient concentration (75.0-96.1%) of PAHs was obtained under the optimised conditions. The method exhibited good reproducibility (RSD values that ranged from 1.5 to 9.0%). The GP-MSE concentration method enhances the volume reduction (concentration factor), leading to a low method detection limit ($0.5-15ngL^{-1}$). Furthermore, this method offers the advantage of small-volume sampling, enabling even the detection of diurnal hourly changes in the concentration of PAHs in ambient air. Utilising this method in combination with GC-MS, the diurnal hourly flux of PAHs from the gas phase of ambient air was measured. Indeed, the proposed technique is a simple, fast, low-cost and environmentally friendly.

Comparative Genomic Analysis and Rapid Molecular Detection of Xanthomonas euvesicatoria Using Unique ATP-Dependent DNA Helicase recQ, hrpB1, and hrpB2 Genes Isolated from Physalis pubescens in China

  • Faisal Siddique;Yang Mingxiu;Xu Xiaofeng;Ni Zhe;Haseeb Younis;Peng Lili;Zhang Junhua
    • The Plant Pathology Journal
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    • 제39권2호
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    • pp.191-206
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    • 2023
  • Ground cherry (Physalis pubescens) is the most prominent species in the Solanaceae family due to its nutritional content, and prospective health advantages. It is grown all over the world, but notably in northern China. In 2019 firstly bacterial leaf spot (BLS) disease was identified on P. pubescens in China that caused by both BLS pathogens Xanthomonas euvesicatoria pv. euvesicatoria resulted in substantial monetary losses. Here, we compared whole genome sequences of X. euvesicatoria to other Xanthomonas species that caused BLS diseases for high similarities and dissimilarities in genomic sequences through average nucleotide identity (ANI) and BLAST comparison. Molecular techniques and phylogenetic trees were adopted to detect X. euvesicatoria on P. pubescens using recQ, hrpB1, and hrpB2 genes for efficient and precise identification. For rapid molecular detection of X. euvesicatoria, loop-mediated isothermal amplification, polymerase chain reaction (PCR), and real-time PCR techniques were used. Whole genome comparison results showed that the genome of X. euvesicatoria was more closely relative to X. perforans than X. vesicatoria, and X. gardneri with 98%, 84%, and 86% ANI, respectively. All infected leaves of P. pubescens found positive amplification, and negative controls did not show amplification. The findings of evolutionary history revealed that isolated strains XeC10RQ, XeH9RQ, XeA10RQ, and XeB10RQ that originated from China were closely relative and highly homologous to the X. euvesicatoria. This research provides information to researchers on genomic variation in BLS pathogens, and further molecular evolution and identification of X. euvesicatoria using the unique target recQ gene through advance molecular approaches.

Enhanced UV-Visible Absorbance Detection in Capillary Electrophoresis Using Modified T-Shaped Post-Column Flow Cell

  • Lim, Kwan-Seop;Kim, Su-Hyeon;Hahn, Jong-Hoon
    • Bulletin of the Korean Chemical Society
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    • 제23권2호
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    • pp.295-300
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    • 2002
  • The construction of the T-shaped post-column flow cell has been changed to enhance the practicability as a UV-visible absorbance detector for capillary electrophoresis. In this new design, a rectangular cube-shaped inner structure is employed, which completely fits the outer rectangular tubing. This arrangement has greatly facilitated the fabrication of the T-cells. In addition, the volume for the auxiliary flow has been dramatically reduced down to 300 ${\mu}L$, and its volume flow rate is optimized at 4.2 ${\mu}L$/min. The short optical path length in the sheath flows (500 ${\mu}m$ on each side) minimizes background absorption, and thus enhances its performance in low-UV wavelengths. We have optimized the auxiliary flow rate at 50 ${\mu}m$/s, so that migration times are insensitive to the flow rate. This optimization has improved repeatabilities in migration times and peak heights. A double-beam detection scheme using a pair of photodiodes is employed to increase the signal-to-noise ratio.

A New Analytical Method to Determine the Purity of Synthetic Fluorophores using Single Molecule Detection Technique

  • Song, Nam-Yoong;Kim, Hyong-Ha;Park, Tae-Sook;Yoon, Min-Joong
    • Journal of Photoscience
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    • 제12권2호
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    • pp.87-93
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    • 2005
  • A new assay technique to distinguish between pure compounds and the isomeric mixtures has been suggested using single molecule (SM) fluorescence detection technique. Since the number of emission spots in a fluorophorespread film prepared from a genuine dye solution was determined by experimental condition, the deviation of spot numbers from the expected values could be considered to be an indication of lower purity of the sample solution. The lower limit of sample concentration for this assay was determined to be $5{\times}10^{-10}$ M to show uniform number of expected spots within 10% uncertainties in our experimental condition. An individual fluorescence intensity distribution for a mixture of isomers having doubly different emissivities was simulated by adding distributions obtained from Cy3 and nile red (NR) independently. The result indicated that the mixture could be identified from the pure compounds through the difference in the number of Gaussian functions to fit the distribution. This new assay technique can be applied to the purity test for synthetic biofluorophores which are usually prepared in small quantities not enough for classical ensemble assays.

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Detection of Pectobacterium chrysanthemi Using Specific PCR Primers Designed from the 16S-23S rRNA Intergenic Spacer Region

  • Kwon, Soon-Wo;Myung, In-Sik;Go, Seung-Joo
    • The Plant Pathology Journal
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    • 제16권5호
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    • pp.252-256
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    • 2000
  • The 16S-23S rRNA intergenic spacer regions (ISRs) were sequenced and analyzed to design specific primer for identification of Pectobacterium chrysanthemi. Two types ISRs, large and small ISRs, were identified from three strains (ATCC 11663, KACC 10163 and KACC 10165) of P. chrysanthemi and Pectobacterium carotovorum subsp. carotovorum ATCC 15713.Large ISRs contained transfer RNA-Ile(tRNA$^{Ile}$)and tRNA$^{Ala}$, and small ISRs contained tRNA$^{Glu}$. Size of the small ISRs of P. chrysanthemi ranged on 354-356 bp, while it was 451 bp in small ISR of P. carotovorum subsp. carotovorum ATCC 15713. From hypervariable region of small ISRs, species-specific primer for P. chrysanthemi with 20 bp length (CHPG) was designed from hypervariable region of small ISRs, which was used as forward promer to detect P. chrysanthemi strains with R23-1R produced PCR product of about 260bp size (CHSF) only from P. chrysanthemi strains, not from other Pectobacterium spp. and Erwinia spp. Direct PCR from bacterial cell without extracting DNA successfully amplified a specific fragment, CHSF, from P. chrysanthemi ATCC 11663. The limit of PCR detection was 1${\pm}10^2$ cfu/ml.

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Molecular Detection of Verotoxigenic Escherichia coli (VTEC) from Animal Feces for Screening VTEC-shedders

  • Kobayashi, Y.;Sato, M.;Taguchi, H.;Koike, S.;Nakatsuji, H.;Tanaka, K.
    • Asian-Australasian Journal of Animal Sciences
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    • 제17권3호
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    • pp.423-427
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    • 2004
  • Seventy-six animals including cattle, sheep, horses, 6 species of zoo animals were employed for collection of fresh feces in order to detect verotoxigenic Esherichia coli (VTEC) by safe, quick and sensitive PCR-based molecular methods. Bacterial cell disruption with bead-beating followed by bacterial DNA purification with hydroxyapatide chromatography and gel filtration allowed DNA preparation from animal feces with high recovery and purity. A mountain goat was firstly shown by PCR and sequencing to shed verotoxin 2 gene (vt2) that was used to generate vt2 probe and second primer set for nested PCR to attempt more sensitive detection. Most sensitive nested PCR revealed that 45% of tested cattle and 47% of tested zoo animals were VTEC-positive, while least sensitive normal PCR detected VTEC from none of these animals except a mountain goat. Moderately sensitive detection by PCR in combination with hybridization suggested that the VTEC density varied between the VTEC-positive cattle.