• Title/Summary/Keyword: real-time polymerase chain reaction

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Combined Detection of Serum MiR-221-3p and MiR-122-5p Expression in Diagnosis and Prognosis of Gastric Cancer

  • Zhang, Yan;Huang, Huifeng;Zhang, Yun;Liao, Nansheng
    • Journal of Gastric Cancer
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    • v.19 no.3
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    • pp.315-328
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    • 2019
  • Purpose: To investigate the clinical value of serum miR-221-3p and miR-122-5p expression levels in the diagnosis and prognosis of gastric cancer. Materials and Methods: Serum samples from 141 gastric cancer cases (gastric cancer group), 110 gastric polyps (gastric polyp group), and 75 healthy people (healthy control) were used to detect miR-221-3p and miR-122-5p expression using real-time reverse transcription polymerase chain reaction. Results: Serum miR-221-3p expression was significantly higher in the gastric cancer group than in the gastric polyp group, and it was significantly lower than that before operation. The miR-221-3p expression was significantly higher in the death group than in the survival group. The proliferation and migration ability significantly increased and the apoptosis rate significantly decreased by miR-221-3p transfection in gastric cancer cells. In contrast, the function of miR-122-5p in gastric cancer cells was opposite of miR-221-3p. Serum miR-221-3p expression was negatively correlated with that of miR-122-5p in gastric cancer. Serum miR-221-3p and miR-122-5p expressions were significantly correlated with the degree of differentiation, tumor, node, metastasis stage, lymph node metastasis, and invasion depth. miR-221-3p and miR-122-5p expression levels were independent prognostic factors for postoperative gastric cancer. In the diagnosis and predicting prognosis of gastric cancer, receiver operating characteristic analysis revealed that the area under curve of combined detection of serum miR-221-3p and miR-122-5p expression had a greater diagnostic effect than either single maker. Conclusions: The miR-221-3p and miR-122-5p are involved in the development of gastric cancer, and they have important clinical values in gastric cancer diagnosis and prognosis.

Discovery of a new primer set for detection and quantification of Ilyonectria mors-panacis in soils for ginseng cultivation

  • Farh, Mohamed El-Agamy;Han, Jeong A.;Kim, Yeon-Ju;Kim, Jae Chun;Singh, Priyanka;Yang, Deok-Chun
    • Journal of Ginseng Research
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    • v.43 no.1
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    • pp.1-9
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    • 2019
  • Background: Korean ginseng is an important cash crop in Asian countries. However, plant yield is reduced by pathogens. Among the Ilyonectria radicicola-species complex, I. mors-panacis is responsible for root-rot and replant failure of ginseng in Asia. The development of new methods to reveal the existence of the pathogen before cultivation is started is essential. Therefore, a quantitative real-time polymerase chain reaction method was developed to detect and quantify the pathogen in ginseng soils. Methods: In this study, a species-specific histone H3 primer set was developed for the quantification of I. mors-panacis. The primer set was used on DNA from other microbes to evaluate its sensitivity and selectivity for I. mors-panacis DNA. Sterilized soil samples artificially infected with the pathogen at different concentrations were used to evaluate the ability of the primer set to detect the pathogen population in the soil DNA. Finally, the pathogen was quantified in many natural soil samples. Results: The designed primer set was found to be sensitive and selective for I. mors-panacis DNA. In artificially infected sterilized soil samples, using quantitative real-time polymerase chain reaction the estimated amount of template was positively correlated with the pathogen concentration in soil samples ($R^2=0.95$), disease severity index ($R^2=0.99$), and colony-forming units ($R^2=0.87$). In natural soils, the pathogen was recorded in most fields producing bad yields at a range of $5.82{\pm}2.35pg/g$ to $892.34{\pm}103.70pg/g$ of soil. Conclusion: According to these results, the proposed primer set is applicable for estimating soil quality before ginseng cultivation. This will contribute to disease management and crop protection in the future.

Comparative Study of Target Genes and Protocols by Country for Detection of SARS-CoV-2 based on Polymerase Chain Reaction (PCR) (중합효소 연쇄반응 기반의 코로나-19 바이러스 검출법에 대한 국가별 목표 유전자 및 프로토콜 비교 연구)

  • Kim, Jin-Hee
    • The Journal of the Korea Contents Association
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    • v.21 no.1
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    • pp.465-474
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    • 2021
  • Corona-19, a disease caused by 'Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2)', was declared a global pandemic by the World Health Organization (WHO) in March 2020, and a real-time polymerase chain reaction test is performed as a diagnostic test for screening and confirmation in most countries. However, not only the target genes and protocols differ by countries, but also the procedures for reading the diagnosis results are diverse, so the criteria for confirmed patients differ by country. Therefore, in this review, we discussed the target genes, test techniques, and diagnostic criteria for each country notified by WHO. And the specificity and sensitivity, limits of detection, positive and negative controls, false positive bacteria candidates, and specimens, and the specifics of the control setting were also described. In addition, the characteristics of Korea's test were compared to each country's one. Finally, in order to obtain the same diagnosis result for SARS-CoV-2 in the future, standardized diagnosis methods and result interpretations for Corona-19 diagnosis were proposed.

Expressional Evaluation of C/EBP Family, SREBP1, and Steroid Hormone Receptors in the Epididiymal Fat of Postnatally Developing Mouse

  • Lee, Yong-Seung;Lee, Ki-Ho
    • Development and Reproduction
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    • v.26 no.2
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    • pp.49-58
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    • 2022
  • The differentiation and development of preadipocyte into mature adipocyte are regulated by transcription factors, such as CCAAT enhancer binding protein (Cebp) gene family and sterol regulatory element binding transcription factor 1 (Srebp1). Steroid hormones give influences on the development and function of adipocyte. The present research examined expression patterns of CCAAT enhancer binding protein alpha (Cebpa), CCAAT enhancer binding protein beta (Cebpb), CCAAT enhancer binding protein gamma (Cebpg), sterol regulatory element binding transcription factor 1 (Srebp1), androgen receptor (Ar), and estrogen receptors (Esr) among different epididymal fat parts during postnatal period by quantitative real-time polymerase chain reaction. In the distal epididymal fat, expression of Cebpa, Cebpb, Cebpg, Srebp1, Ar, and Esr2 was increased until 12 months of age, while expression of Esr1 was decreased at 5 months of age and was not detectable after 8 months of age. In the proximal epididymal fat, transcript levels of Cebps and Srebp1 were increased at 8 months of age, followed by decreases of Cebpb and Cebpg transcript levels at 12 months of age. An additional increase of Srebp1 expression was observed at 12 months of age. Expression of Ar and Esr2 were increased until 8 months of age, followed by a drop of Ar expression level at 12 months of age. Expression pattern of Esr1 was similar to that in the distal epididymal fat. In the tail epididymal fat, expression of Cebpa, Cebpg, Srebp1, Ar, and Esr2 was increased with age. Esr1 was not detectable at all. The highest level of Cebpb was observed at 8 months of age. These data suggest the possibility of developmental and functional differentiation among the epididymal fat parts.

Effect of Tetrodotoxin on the Proliferation and Gene Expression of Human SW620 Colorectal Cancer Cells

  • Bae, Yun-Ho;Kim, Hun;Lee, Sung-Jin
    • Biomedical Science Letters
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    • v.28 no.1
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    • pp.42-49
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    • 2022
  • Tetrodotoxin (TTX) is a natural neurotoxin found in several species of puffer fish belonging to Tetraodon fugu genus and has been reported to affect processes such as proliferation, metastasis and invasion of various cancer cells. However, it was not revealed which genes were influenced by these reactions. In this experiment, it was examined in human SW620 colorectal cancer cells. The proliferation of SW620 cells was significantly reduced when treated with 0, 1, 10 and 100 μM TTX for 48 h. It was confirmed using Annexin V-propidium iodide staining that some apoptosis was induced. Differentially expressed genes (DEGs) affecting cell proliferation through RNA sequencing (RNA-seq) were selected. The expression change of DEGs was confirmed by conducting quantitative real-time polymerase chain reaction (qRT-PCR). As a result, the mRNA expression of FOS and WDR48 genes was found to be increased in the 100 μM TTX treatment group compared to the control group. On the other hand, the mRNA expression of ALKBH7, NDUFA13, RIPPLY3 and SELENOM genes was found to be reduced, and in the case of the ALKBH7 gene was identified to show significant differences. This experiment suggests that TTX can be used as an important fundamental data to elucidate the mechanism that inhibits the proliferation of SW620 cells.

Applying nano-HA in addition to scaling and root planing increases clinical attachment gain

  • Uysal, Ozge;Ustaoglu, Gulbahar;Behcet, Mustafa;Albayrak, Onder;Tunali, Mustafa
    • Journal of Periodontal and Implant Science
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    • v.52 no.2
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    • pp.116-126
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    • 2022
  • Purpose: This study evaluated the efficacy of treating periodontitis using subgingival nano-hydroxyapatite powder with an air abrasion device (NHAPA) combined with scaling and root planing (SRP). Methods: A total of 28 patients with stage III periodontitis (grade B) were included in this study, although 1 was lost during follow-up and 3 used antibiotics. The patients were divided into a test group and a control group. All patients first received whole-mouth SRP using hand instruments, and a split-mouth approach was used for the second treatment. In the test group, the teeth were treated with NHAPA for 15 seconds at 70% power per pocket. Subgingival plaque samples were obtained from the 2 deepest pockets at the test and control sites before treatment (baseline) and 3 months after treatment. The full-mouth plaque index (PI), gingival index (GI), papillary bleeding index (PBI), bleeding on probing (BOP), probing depth (PD) and clinical attachment level (CAL) were recorded at baseline and at 1- and 3-month post-treatment. Real-time polymerase chain reaction was used to determine the colonisation of Treponema denticola (Td), Porphyromonas gingivalis (Pg), and Aggregatibacter actinomycetemcomitans in the subgingival plaque. Results: From baseline to the first month, the test group showed significantly larger changes in BOP and CAL (43.705%±27.495% and 1.160±0.747 mm, respectively) than the control group (36.311%±27.599% and 0.947±0.635 mm, respectively). Periodontal parameters had improved in both groups at 3 months. The reductions of PI, GI, BOP, PD, and CAL in the test group at 3 months were greater and statistically significant. The total bacterial count and Td and Pg species had decreased significantly by the third month in both groups (P<0.05). Conclusions: Applying NHAPA in addition to SRP improves clinical periodontal parameters more than SRP alone. Subgingival NHAPA may encourage clot adhesion to tooth surfaces by increasing surface wettability.

Assessment of Adipocyte Differentiation and Maturation-related Gene Expression in the Epididymal Fat of Estrogen Receptor α Knockout (ERαKO) Mouse during Postnatal Development Period

  • Cheon, Yong-Pil;Ko, CheMyong;Lee, Ki-Ho
    • Development and Reproduction
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    • v.24 no.4
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    • pp.287-296
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    • 2020
  • The absence of functional estrogen receptor α (Esr1) results in an overgrowth of the epididymal fat, as observed in estrogen receptor α knockout (ERαKO) mouse. The present research was aimed to evaluate expression of various molecules associated with adipocyte differentiation and maturation in the epididymal fat of ERαKO mouse at several postnatal ages by using quantitative real-time polymerase chain reaction. The highest transcript levels of all molecules were detected at 12 months of postnatal age, except leptin which the mRNA level was increased at 5 months of age and was unchanged until 12 months of age. The expression levels of CCAAT enhancer binding protein (Cebp) alpha, androgen receptor, and lipoprotein lipase were decreased at 5 months of age but increased at about 8 months of age. The mRNA levels of Cebp gamma and sterol regulatory element binding transcription factor 1 remained steady until 8 months of age. Continuous increases of transcript levels during postnatal period were found in Cebp beta, estrogen receptor (ER) beta, fatty acid binding protein 4, and delta like non-canonical Notch ligand 1. The increases of peroxisome proliferator-activated receptor gamma and adiponectin mRNA levels were detected as early as 8 months of age. The levels of fatty acid synthase and resistin transcript at 5 and 8 months of age were lower than that at 2 months of age. These findings show the aberrant expression patterns of genes related to adipocyte differentiation and maturation in the postnatal epididymal fat pad by the disruption of ER alpha function.

Comparison of 10 Different Pre-Enrichment Broths for the Regeneration of Cronobacter spp. (Enterobacter sakazakii ) Infected in Powdered Infant Formula

  • Jung-Whan Chon;Kun-Ho Seo;Hyungsuk Oh;Dongkwan Jeong;Kwang-Young Song
    • Journal of Dairy Science and Biotechnology
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    • v.41 no.3
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    • pp.103-112
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    • 2023
  • This study aimed to assess the effectiveness of 10 different pre-enrichment methods using Real-Time polymerase chain reaction (PCR) in support of the FDA method. When the initial Cronobacter spp. (Enterobacter sakazakii) inoculation was 7.2 CFU/g, the Ct values were observed in the following order: 21.37 (Enterobacteriaceae enrichment [EE] broth), 21.95 (brain heart infusion [BHI]), 22.72 (tryptic soy broth [TSB]), 23.02 (violet red bile lactose [VRBL]), 22.31 (TSB-0.1% sodium pyruvate [SP]), 23.43 (distilled water [DW]), 24.34 (phosphate buffered saline [PBS]), 24.95 (nutrient broth [NB]), 25.82 (TSB-0.6% yeast extract [YE]), and 28.27 (violet red bile glucose [VRBG]). For an inoculation of 1.82% CFU/g of Cronobacter spp. (E. sakazakii), the Ct values were recorded in this sequence: 20.34 (EE broth), 22.16 (TSB-0.6% YE), 22.37 (BHI), 22.71 (VRBL), 22.88 (TSB), 23.01 (DW), 23.19 (NB), 23.79 (TSB-0.1% SP), 24.66 (VRBG), and 24.70 (PBS). Finally, when the inoculum of Cronobacter spp. (E. sakazakii) was 0.182 CFU/g, the Ct values followed this order: 21.93 (VRBL), 23.07 (TSB-0.6% YE), 23.31 (DW), 23.47 (PBS), 23.70 (BHI), 24.14 (TSB-0.1% SP), 25.14 (TSB), 29.00 (VRBG), 31.55 (EE broth), and were undetected in the case of NB. Consequently, these results indicate that there were no significant differences among the 10 different pre-enrichment broths. Future studies should focus on exploring pre-enrichment broths that can improve the limit of detection at very low Cronobacter spp. (E. sakazakii) concentrations and enhance the selective recovery of Cronobacter spp. (E. sakazakii) under acid, antibiotic, cold, and heat damage conditions.

MALDI TOF MS for the identification of Salmonella spp. from swine (돼지유래 Salmonella속 균의 동정을 위한 MALDI TOF MS 활용)

  • Sohn, Jun Hyung;Jeon, Woo Jin;Lee, Young Mi;Kim, Seon Soo
    • Korean Journal of Veterinary Service
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    • v.39 no.4
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    • pp.247-251
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    • 2016
  • Salmonella is one of the most common bacteria that causes heavy losses in swine industry and major causative pathogen of food poisoning in public health. Various methods for the identification of Salmonella such as Gram staining, agglutination test, enzyme-linked immunosorbent assay (ELISA), polymerase chain reaction (PCR) have been used. Several studies have demonstrated that Matrix Assisted Laser Desorption Ionization Time of Flight (MALDI TOF) Mass Spectrometry (MS) identification is an efficient and inexpensive method for the rapid and routine identification of isolated bacteria. In this study, MALDI TOF MS could provide rapid, accurate identification of Salmonella spp. from swine compared with end point PCR and real time PCR.

Epidemiology and Characteristics of Pediatric Respiratory Virus Infection From 2017 to 2019 Focusing on Human Coronavirus: A Retrospective Study of a Single Center in Northwestern Gyeonggi-do (인간 코로나 바이러스를 중심으로 2017-2019년 소아청소년 호흡기 바이러스 감염증의 역학 및 특성: 경기 북서부지역 단일기관의 후향적 연구)

  • Hyoungsuk Park;Kyoung Won Cho;Lindsey Yoojin Chung;Jong Min Kim;Jun Hyuk Song;Kwang Nam Kim
    • Pediatric Infection and Vaccine
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    • v.30 no.2
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    • pp.62-72
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    • 2023
  • Purpose: A change is expected in the pattern of respiratory viruses including human coronavirus (HCoV) after the coronavirus disease 2019 (COVID-19) outbreak. Accordingly, identifying the distribution of respiratory viruses before the COVID-19 outbreak is necessary. Methods: We retrospectively analyzed the results of samples of nasal swabs collected from children under aged ≤18 years who were hospitalized at Myongji Hospital, Gyeonggi-do due to acute respiratory infections from 2017 to 2019. Viruses were detected by real-time reverse transcription polymerase chain reaction (RT-PCR). Results: Out of 3,557 total patients, 3,686 viruses were detected with RT-PCR including coinfections. Of the 3,557 patients, 2,797 (78.6%) were confirmed as PCR-positive. Adenovirus and human rhinovirus (hRV) were detected throughout the year, and human enterovirus was most detected during summer. Respiratory syncytial virus, influenza virus, and HCoV were prevalent in winter. In patients with croup, parainfluenza virus was most frequently detected, followed by hRV and HCoV. The PCR positive rate in summer and winter differed significantly. Conclusions: Respiratory virus patterns in northwestern Gyeonggi-do were not much different from previously reported data. The data reported herein regarding respiratory virus epidemiological information before the COVID-19 outbreak can be used for use in comparative studies of respiratory virus patterns after the COVID-19 outbreak.