• Tuberculosis and Respiratory Diseases
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• v.49 no.5
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• pp.546-557
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• 2000

Background : Oligonucleotide chip technology has proven to be a very useful tool in the rapid diagnosis of infectious disease. Rifampin resistance is considered as a useful marker of multidrug-resistance in tuberculosis. Mutations in the rpoB gene coding $\beta$ subunit of RNA polymerase represent the main mechanism of rifampin resistance. The purpose of this study was to develop a diagnosis kit using oligonucleotide chip for the rapid and accurate detection of rifampin-resistance in Mycobacterium tuberculosis. Method : The sequence specific probes for mutations in the rpoB gene were designed and spotted onto the glass slide, oligonucleotide chip. 38 clinical isolates of Mycobacterium were tested. A part of rpoB was amplified, labelled, and hybridized on the oligonucleotide chip with probes. Results were analyzed with a laser scanner. Direct sequencing was done to verify the results. Result : The low-density oligonucleotide chip design어 to determine the specific mutations in the rpoB gene of M. tuberculosis accurately detected rifampin resistance associated with mutations in 28 clinical isolates. Mutations at codons 531, 526, and 513 were confirmed by direct sequencing analysis. Conclusion : Mutant detection using oligonucleotide chip technology is a reliable and useful diagnostic tool for the detection of multidrug-resistance in M. tuberculosis.

• Proceedings of the Korean Society of Fisheries Technology Conference
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• 2003.05a
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• pp.309-310
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• 2003

현재 어류 질병 바이러스를 검출해내기 위해 이용되는 PCR법이나 ELIZA법 만으로는 신속성을 필요로 하는 바이러스 검출에서는 불리하다(Bruchhof et. al., 1995). 따라서 기존의 유전자 진단법을 대체할 수 있고, 동시에 수백 개 이상의 유전자를 빠른 시간 내에 검색할 수 있는 DNA chip 기술을 이용하여 넙치, 돔등의 해산어와 일부 연어과 어류에 질병을 일으키는 rhabdovirus를 신속하고 정확하게 진단할 수 있는 저밀도 oligonucleotide chip을 개발하고자 한다(Chizhikov et at., 2001). (중략)

• Proceedings of the Korean Society of Fisheries Technology Conference
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• 2003.05a
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• pp.311-312
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• 2003

Microarray를 해석하는데 있어서 방해가 되는 요인에는 여러 가지가 있을 수 있다. 예를 들면 probe DNA의 농도, spotting 시의 습도, spot의 크기 및 모양, slide blocking 과정, lab리ing technique, hybri야zation 온도와 시간 그리고 background signal 등이 그것이다(Hegde et at., 2000). 본 연구에서는 전보에서 선정된 rhabdovirus의 96개 probe로 실제 oligonucleotide chip을 만들어 spot의 모양이나 크기에 관계되는 spotting 조건을 확립하고, slide blocking과 slide washing에 따른 background signal을 낮추기 위한 과정을 도입하였다. (중략)

• Tuberculosis and Respiratory Diseases
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• v.50 no.1
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• pp.29-41
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• 2001

Background : The appearance of multiple-drug-resistant Mycobacterium tuberculosis strains has been seriously compromising successful control of tuberculosis. Rifampin-resistance, caused by mutations in the rpoB gene, can be indicative of multiple-drug-resistance, and its detection is of great importance. The present study aimed to develop an oligonucleotide chip for accurate and convenient screening of drug-resistance. Methods : In order to detect point mutations in the rpoB gene, an oligonucleotide chip was prepared by immobilizing specific probe DNA to a microscopic slide glass by a chemical reaction. The probe DNA that was selected from the 81 bp core region of the rpoB gene was designed to have mutation sites at the center. A total of 17 mutant probes related to rifampin-resistance including 8 rifabutin-sensitive mutant probes were used in this study. For accurate determination, wild type probes were prepared for each mutation position with an equal length, which enabled a direct comparison of the hybridization intensities between the mutant and wild type. Results : Mycobacterial genomic DNA from clinical samples was tested with the oligonucleotide chip and the results were compared with those of the drug-susceptibility test in addition to sequencing and INNO-LiPA Rif. TB kit test in some cases. Out of 15 samples, the oligonucleotide chip results of 13 samples showed good agreement with the rifabutin-sensitivity results. The two samples with conflicting result also showed a discrepancy between the other tests, suggesting such possibilities as existence of mixed strains and difference in drug-sensitivity. Further verification of these samples in addition to more case studies are required before the final evaluation of the oligonucleotide chip can be made. Conlcusion : An oligonucleotide chip was developed for the detection of rpoB gene mutations related to drugsusceptibility. The results to date show the potential for using the oligonucleotide chip for accurate and convenient screening of drug-resistance to provide useful information in antituberculosis drug therapy.

• The Plant Pathology Journal
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• v.30 no.1
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• pp.51-57
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• 2014

A large-scale oligonucleotide (LSON) chip was developed for the detection of the plant viruses with known genetic information. The LSON chip contains two sets of 3,978 probes for 538 species of targets including plant viruses, satellite RNAs and viroids. A hundred forty thousand probes, consisting of isolate-, species- and genus-specific probes respectively, are designed from 20,000 of independent nucleotide sequence of plant viruses. Based on the economic importance, the amount of genome information, and the number of strains and/or isolates, one to fifty-one probes for each target virus are selected and spotted on the chip. The standard and field samples for the analysis of the LSON chip have been prepared and tested by RT-PCR. The probe's specific and/or nonspecific reaction patterns by LSON chip allow us to diagnose the unidentified viruses. Thus, the LSON chip in this study could be highly useful for the detection of unexpected plant viruses, the monitoring of emerging viruses and the fluctuation of the population of major viruses in each plant.

• Tuberculosis and Respiratory Diseases
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• v.55 no.1
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• pp.41-58
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• 2003

Background : The resurgence of tuberculosis and the widespread emergence of multidrug-resistant M. tuberculosis have emphasized the importance of rapid and accurate diagnostic procedures. Recently, the oligonucleotide chip has proven to be a useful tool in the rapid diagnosis of infectious diseases. The purpose of this study was to rapidly and accurately detect specific mutations in the rpoB, katG and rpsL genes associated with rifampin, isoniazid and streptomycin resistance in M. tuberculosis, respectively, using a single oligonucleotide chip. Method : For detection of drug-resistance, 7 wild-type and 13 mutant-type probes for rifampin, 2 wild-type and 3 mutant-type probes for isoniazid, and 2 wild-type and 2 mutant-type probes for streptomycin were designed and spotted onto glass slides. Fifty-five cultured samples of M. tuberculosis were amplified by PCR, and then underwent hybridization and scanning. Direct sequencing was done to verify the results from the oligonucleotide chip and to analyze the types of mutations. Result : Thirty-five cases out of 40 rifampin-resistant strains(~88%) had mutations in the rpoB gene. One case had a new mutation(D516F, GAC R TTC) and another known mutation together. Twenty cases out of 42 isoniazid-resistant strains(~50%) had mutations in the katG gene, while 7 cases out of 9 streptomycin-resistant strains(~78%) had mutations in the rpsL gene. From these results, the oligonucleotide chip was confirmed to be able to detect the most frequent mutations from the genes associated with rifampin, isoniazid and streptomycin resistance. The results proved that the drug-resistance detection probes were specific. When the results from the oligonucleotide chip and DNA sequencing were compared, the types of mutations were exactly matched. Conclusion : The diagnostic oligonucleotide chip with mutation specific probes for drug resistance is a very reliable and useful tool for the rapid and accurate diagnosis of drug resistance against rifampin, isoniazid and streptomycin in M. tuberculosis infections.

• Korean Journal of Acupuncture
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• v.23 no.2
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• pp.125-136
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• 2006

Objectives: It has long been known about the osteogenic effect of CPC-HAS(cervi parvum cornu herbal-acupuncture solution) on bone tissues. However, it has not been determined the effect of CPC-HAS on cancer cells. The purpose of this study is to screen the CPC-HAS mediated differentially expressed genes..in cancer cells such as SNU484 gastric cancer cell lines. Oligonucleotide microarray approache was employed to screen the differential expression genes. Methods: CPC-HAS was prepared by boiling and stored at $-70^{\circ}C$ until use. Cells were treated with various concentrations of CPC-HAS (0.1, 0.5, 1.5, 10, 20 mg/ml) for 24 h. Cell toxicity was tested by MTT assay. To screen the differentially expressed genes in cancer cells, cells were treated with 1.5 mg/ml of CPC-HAS. For oligonucleotide microarray assay, total RNA was used for gene expression analysis using oligonucleotide Genechip(Human genome U133 Plus 2.0., Affimatrix Co.). Results: It has no cytotoxic effects on SNU484 cell in all concentrations(0.l, 0.5, 1.5, 10, 20 mg/ml). In oligonucleotide microarray assay, in SNU484 cells, the number of more than twofold up-regulated genes was 5 while, the number of more than twofold down-regulated genes was 10. Conclusions: This study showed the screening of CPC-HAS mediated differentially regulated genes using combined approaches of oligonucleotide microarray. The screened genes will be used for the better understanding of the therapeutic effects of CPC-HAS on cancer fields.

• Korean Journal of Veterinary Service
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• v.30 no.4
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• pp.489-496
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• 2007

An oligonucleotide microarray system was developed for the simultaneous detection of porcine epidemic diarrhea virus, transmissible gastroenteritis virus, porcine enteric calicivirus, porcine group A and C rotavirus. RNAs of the reference viruses and porcine diarrhea samples were extracted and amplified using one-step multiplex RT-PCR in the presence of cyanine 5-dCTP and hybridized on the microarray chip that spotted the virus-specific oligonucleotides. This system were approximately 10-to 100-fold higher in sensitivity than conventional RT-PCR, and the assay time was less than 3 hours. The relative sensitivity and specificity were 92% and 72.2%, respectively, based on 102 porcine diarrhea samples using RT-PCR as gold standard. These results suggested that the oligonucleotide microarray system in this study be probably more reliable and reproducible means for detecting porcine enteric viruses and that it could be of substantial use in routine diagnostic laboratories.

• KIEE International Transactions on Electrophysics and Applications
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• v.11C no.3
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• pp.85-90
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• 2001

The determination of DNA hybridization reaction can apply the molecular biology research, clinic diagnostics, bioengineering, environment monitoring, food science and other application area. So, the improvement of DNA detection system is very important for the determination of this hybridization reaction. In this study, we report the characterization of the probe and target oligonucleotide hybridization reaction using the evanescent field microscopy. First, we have fabricated DNA chip microarray. The particles which were immobilized oligonucleotides were arranged by the random fluidic self-assembly on the pattern chips, using hydrophobic interaction. Second, we have detected DNA hybridization reaction using evanescent field microscopy. The 5'-biotinylated probe oligonucleotides were immobilized on the surface of DNA chip microarray and the hybridization reaction with the Rhodamine conjugated target oligonucleotide was excited fluorescence generated on the evanescent field microscopy. In the foundation of this result, we could be employed as the basis of a probe olidonucleotide, capable of detecting the target oligonucleotide and monitoring it in a large analyte concentration range and various mismatching condition.

• Korean Journal of Microbiology
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• v.38 no.3
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• pp.186-191
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• 2002

The usefulness of oligonucleotide DNA chip was evaluated for detection and primary level identification of major waterborne viruses in environmental samples. The enteric waterborne viruses included enterovirus, adenovirus, and rotavirus. Total intracellular RNA of 10 BGM cell plates showing virus-specific cytopathic effects was extracted at the third day after inoculation. The intracellular RNA was then subjected to either enterovirus-specific RT-PCR followed by sequencing analysis, or the DNA chip. Seven out of 10 positive samples in cell culture were positive but the other three sample were turned out to be negative by both RT-PCR and DNA chip analyses. Nucleotide sequencing results and the DNA chip hybridization results of the RT-PCR product were in complete agreement in the identification of the 7 positive samples as enteroviruses. Using the DNA chip, it took only 3∼4 hr to complete detection and primary level identification of target viruses and additional procedures such as gel electrophoresis or nucleotide sequencing were not necessary. We believe that the DNA chip system can be employed as a highly effective and new detection methodology for environmental viruses.