• Journal of Animal Science and Technology
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• 제46권5호
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• pp.841-848
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• 2004

살균된 우유에 오염된 Salmonella의 신속한 검출 방법을 공중 보건의 보호를 위하여 중요하다. RT-PCR 방법은 생균과 사균을 분별하여 검출할 수 있는 분자유전학적 방법이다. 본 연구의 RT-PCR 방법은 Salmonella의 type 1 fImbriae의 단량체 단백질을 암호화하는 fimA 유전자의 mRNA를 주형으로 하여 DNA를 증폭하는 방법으로서 활력이 있는 Salmonella를 검출할수 있기 위하여 개발되었다. RNA 시료를 RQRNase-free DNase로 처리하여 오염된 DNA를 파괴하여 RT-PCR 반응에서 주형으로서 DNA가 합성되는 것을 방지할 수 있었다. 이 RT-PCR 방법은 Salmonella 7균주를 검출하였으나 Escherichia coli, Shigella sonnei, Citrobacter freundii, 및 Klebsiella pneumoniae 는 검출하지 않았다. 우유 내에서 열처리된 $10^7/ml과 10^6/ml$ 세균수의 Salmonella는 검출되 었으나 $10^5{\sim}100/ml$는 검출되지 않았다. RT-PCR를 검출할 수 있는 활력이 있는 Salmonella의 최소 세균수는 100/ml이었다.

• Korean Journal of Radiology
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• 제21권4호
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• pp.505-508
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• 2020

The epidemic of 2019 novel coronavirus, later named as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is still gradually spreading worldwide. The nucleic acid test or genetic sequencing serves as the gold standard method for confirmation of infection, yet several recent studies have reported false-negative results of real-time reverse-transcriptase polymerase chain reaction (rRT-PCR). Here, we report two representative false-negative cases and discuss the supplementary role of clinical data with rRT-PCR, including laboratory examination results and computed tomography features. Coinfection with SARS-COV-2 and other viruses has been discussed as well.

• Molecules and Cells
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• 제23권3호
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• pp.287-303
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• 2007

Expressed sequence tags (ESTs) and differentially expressed cDNAs from the self-fertilizing fish, Kryptolebias marmoratus were mined to develop alternative biomarkers for endocrine-disrupting chemicals (EDCs). 1,577 K. marmoratus cDNA clones were randomly sequenced from the 5'-end. These clones corresponded to 1,518 and 1,519 genes in medaka dbEST and zebrafish dbEST, respectively. Of the matched genes, 197 and 115 genes obtained Unigene IDs in medaka dbEST and zebrafish dbEST, respectively. Many of the annotated genes are potential biomarkers for environmental stresses. In a differential display reverse transcriptase-polymerase chain reaction (DD RT-PCR) study, 56 differential expressed genes were obtained from fish liver exposed to bisphenol A. Of these, 16 genes were identified after BLAST search to GenBank, and the annotated genes were mainly involved in catalytic activity and binding. The expression patterns of these 16 genes were validated by real-time RT-PCR of liver tissue from fish exposed to bisphenol A. Our findings suggest that expression of these 16 genes is modulated by endocrine disrupting chemicals, and therefore that they are potential biomarkers for environmental stress including EDCs exposure.

• 한국영양학회:학술대회논문집
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• 한국영양학회 2003년도 연합학술대회
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• pp.132.2-132.2
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• 2003

• 한국어병학회지
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• 제5권2호
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• pp.143-152
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• 1992

Metallothionein은 세포내의 중금속의 농도을 조절하는 주요한 단백질로서 bacteria에서 척추동물에 이르기까지 모든 생명체에서 나타나는 공통된 단백질이다. 비록 metallothionein의 정확한 기능은 알려져 있지 않으나 독성을 나타내는 중금속에 대하여 세포내 방어기작에 관여할 뿐만 아니라 여러다른 유전자의 총괄적 조절기작 및 matalloprotein의 발현에 관여할 것으로 보고있다. 본 연구에서는 Channel Catfish의 metallothionein cDNA 유전자를 poly(A)를 갖는 mRNA로 부터 Reverse Transcriptase-Polymerase Chain Reaction(RT-PCR)에 의하여 cloning하였다. 증폭된 PCR products는 pBluescript SK+의 EcoRV site 및 pUC19의 Smal site에 dT tailing을 하여 cloning하였으며, PCR products는 multicloning site에 있는 EcoRI 및 HindIII 로 절단하여 확인하거나 신속한 PCR screening에 의하여 확인하였다. 여러 PCR clone 중 하나인 pMT150에 대한 DNA 염기서열을 조사한 결과 다른 어류의 metallothionein cDNA 유전자와 높은 유사성을 보였다.

• Pediatric Infection and Vaccine
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• 제25권2호
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• pp.91-100
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• 2018

목적: 본 연구는 30일 이상 90일 미만의 발열이 있는 영아에서 경험적 항생제 사용에 미치는 요소를 조사하였다. 방법: 2010년 1월부터 2016년 12월까지 부산대학교 어린이병원에 발열이 있는 이전에 건강했던 영아를 대상으로 임상양상, 검사소견, 항생제 사용에 대해 의무기록을 후향적으로 분석하였다. 호흡기 바이러스는 다중 역전사 중합 연쇄반응검사로 검출하여 1-3일 후 보고되었고, enterovirus는 중합 연쇄반응검사로 검출하여 수시간 만에 보고되었다. 결과: 366명의 대상자 중 129명은 경험적 항생제를 사용하였고 237명은 경험적 항생제를 사용하지 않았다. 입원 전 발열기간이 긴 경우와 호흡기 증상이 있을 때, 아파보일 때, CRP 상승 시 경험적 항생제 사용이 많았다. 경험적 항생제를 사용하지 않은 환자의 재입원율이 낮았다. Enterovirus polymerase chain reaction (PCR)이 검출된 대부분의 환자는 경험적 항생제를 사용하지 않았다. Respiratory virus multiplex reverse transcriptase (RT)-PCR 결과는 경험적 항생제 사용에 차이를 보이지 않았다. 결론: 본 연구에서 respiratory virus multiplex RT-PCR은 항생제 처방에 영향을 주지 않았고 enterovirus PCR은 항생제 처방에 영향을 주었다. Respiratory virus multiplex RT-PCR 결과가 신속하게 보고된다면 항생제 사용에 영향을 줄 것이다.

• 한국동물위생학회지
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• 제38권2호
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• pp.107-116
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• 2015

In this study, we developed a rapid, sensitive and specific reverse-transcriptase loop-mediated isothermal amplification (RT-LAMP) assay for detection of swine influenza viruse (SIV) including major subtypes of swine influenza viruses H1N1, H1N2 and H3N2, and a novel subtype of influenza A virus that accidentally infected in pig population. The RT-LAMP was completed in 40 min at $58^{\circ}C$ and the sensitivity of the RT-LAMP ($1copy/{\mu}L$) was 10-fold higher than conventional reverse transcription-polymerase chain reaction (RT-PCR) ($10copy/{\mu}L$) and the same to real time RT-PCR ($1copy/{\mu}L$). Also, the result of the RT-LAMP can be confirmed without any detection system. Therefore, the RT-LAMP could be a alternative diagnostic method for SIV detection in national SIV monitoring system and clinical diagnostic laboratory in the future.

• Archives of Pharmacal Research
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• 제19권6호
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• pp.486-489
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• 1996

A simple, rapid, specific and sensitive method for the detection of serum hepatitis C virus (HCV) RNA using the reverse transcription-polymerase chain reaction (RT-PCR) technique without conventional RNA extraction was developed. HCV template RNA from serum was obtained by boiling the serum at $95^{\circ}C$ for 2 min, cooling rapidly in ice and removing the proteins by cetrifugation. RT-PCR amplifications including the reverse transcription and first PCR amplification were performed in one vessel containing both of reverse transcriptase and Taq DNA polymerase. The detection of HCV RNA from $10^{-3}{\mu}l$. serum was possible with this method. The suitability of this method for clinical analysis was evaluated by assaying HCV RNA in 225 patient samples including anti-HCV antibody negatives (13 samples) and positives (212 samples) by enzyme-linked immunosorbent assay test (ELISA). Detections of HCV RNA with this method were in 4 of 13 anti-HCV antibody negative samples (30.8%) and 95 of 212 positive samples (44.8%). The present method can be completed in 1 hr and has a wide range of application for the clinical utilities to determine the viral RNAS.

• Asian-Australasian Journal of Animal Sciences
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• 제18권6호
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• pp.892-897
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• 2005

Macrophage colony-stimulating factor (M-CSF) is a growth factor required for growth and differentiation of mononuclear phagocyte lineage. Total and 16 poly (A) mRNA of bovine M-CSF were isolated from healthy bovine peripheral mononuclear cells stimulated by phobol 12-myristste 13-acetate (TPA). The more compatible cultured mononuclear cells were 5${\times}$10/ml for RNA isolation. TPA-activated mononuclear cells increased the level of M-CSF-mRNA more than concanavalin A (Con A) and lipopolysaccharide (LPS). The optimal analysis of reverse transcriptase-polymerase chain reaction (RT-PCR) for14 Macrophage colonystimulating factor (M-CSF) as a growth factor required for bovine M-CSF was denaturation at 94$^{\circ}C$ for 1 minute, annealing at 57$^{\circ}C$ for 1 minute, extension at 72$^{\circ}C$ for 1 minute for 30 cycles. The size of cDNA of bovine M-CSF by RT-PCR was 774 base pairs. A 774 base pairs cDNA encoding bovine M-CSF was synthesized by reverse transcriptase polymerase chain reaction (RT-PCR). Ligated cDNA was transformed to competent cells and then plasmid isolation and digestion was performed. Molecular cloning and sequencing were performed for cDNA of bovine M-CSF. The size of cloned cDNA of bovine M-CSF was 774base pairs. The homology of base sequence and amino acid sequence was 88% and 86% compared with known human M-CSF, respectively. From a high degree of sequence similarity, the obtained cDNA of bovine M-CSF is thought be a specific gene of bovine M-CSF.

• Clinical and Experimental Pediatrics
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• 제55권12호
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• pp.470-473
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• 2012

Purpose: The purpose of this prospective case-control study was to survey the detection rate of respiratory viruses in children with Kawasaki disease (KD) by using multiplex reverse transcriptase-polymerase chain reaction (RT-PCR), and to investigate the clinical implications of the prevalence of respiratory viruses during the acute phase of KD. Methods: RT-PCR assays were carried out to screen for the presence of respiratory syncytial virus A and B, adenovirus, rhinovirus, parainfluenza viruses 1 to 4, influenza virus A and B, metapneumovirus, bocavirus, coronavirus OC43/229E and NL63, and enterovirus in nasopharyngeal secretions of 55 KD patients and 78 control subjects. Results: Virus detection rates in KD patients and control subjects were 32.7% and 30.8%, respectively (P=0.811). However, there was no significant association between the presence of any of the 15 viruses and the incidence of KD. Comparisons between the 18 patients with positive RT-PCR results and the other 37 KD patients revealed no significant differences in terms of clinical findings (including the prevalence of incomplete presentation of the disease) and coronary artery diameter. Conclusion: A positive RT-PCR for currently epidemic respiratory viruses should not be used as an evidence against the diagnosis of KD. These viruses were not associated with the incomplete presentation of KD and coronary artery dilatation.