• Title/Summary/Keyword: Direct PCR

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Painless Jaundice Caused by Clonorchis sinensis Infection: A Case Report

  • Hao, Yuhua;Bao, Wanguo;Jin, Meishan;Li, Yuxiang;Wang, Feng
    • Parasites, Hosts and Diseases
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    • v.54 no.3
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    • pp.323-327
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    • 2016
  • A man with only yellowing of the skin and eye sclera was diagnosed with clonorchiasis, which rarely manifested jaundice as the initial symptom. However, because of a lack of evidence for a diagnostic gold standard, the time until definitive diagnosis was more than a week. The diagnostic process relied on inquiring about the patient's history, including the place of residence, dietary habits, and symptoms, as well as on serological findings, an imaging examination, and pathological findings. MRCP and CT results showed mild dilatation of intrahepatic ducts and increased periductal echogenicity. The eggs were ultimately found in stool by water sedimentation method after the negative report through direct smear. DNA sequencing of PCR production of the eggs demonstrated 98-100% homology with ITS2 of Clonorchis sinensis. After anti-parasite medical treatment, the patient's symptoms were gradually relieved. Throughout the diagnostic procedure, besides routine examinations, the sedimentation method or concentration method could be used as a sensitive way for both light and heavy C. sinensis infection in the definite diagnosis.

MiR-150-5p Suppresses Colorectal Cancer Cell Migration and Invasion through Targeting MUC4

  • Wang, Wei-Hua;Chen, Jie;Zhao, Feng;Zhang, Bu-Rong;Yu, Hong-Sheng;Jin, Hai-Ying;Dai, Jin-Hua
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.15
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    • pp.6269-6273
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    • 2014
  • Growing evidence suggests that miR-150-5p has an important role in regulating genesis of various types of cancer. However, the roles and the underlying mechanisms of miR-150-5p in development of colorectal cancer (CRC) remain largely unknown. Transwell chambers were used to analyze effects on cell migration and invasion by miR-150-5p. Quantitative real-time PCR (qRT-PCR), Western blotting and dual-luciferase 3' UTR reporter assay were carried out to identify the target genes of miR-150-5p. In our research, miR-150-5p suppressed CRC cell migration and invasion, and MUC4 was identified as a direct target gene. Its effects were partly blocked by re-expression of MUC4. In conclusiomn, miR-150-5p may suppress CRC metastasis through directly targeting MUC4, highlighting its potential as a novel agent for the treatment of CRC metastasis.

Comparison of Alpha-Factor Preprosequence and a Classical Mammalian Signal Peptide for Secretion of Recombinant Xylanase xynB from Yeast Pichia pastoris

  • He, Zuyong;Huang, Yuankai;Qin, Yufeng;Liu, Zhiguo;Mo, Delin;Cong, Peiqing;Chen, Yaosheng
    • Journal of Microbiology and Biotechnology
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    • v.22 no.4
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    • pp.479-483
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    • 2012
  • The secretory efficiency of recombinant xylanase xynB from yeast Pichia pastoris between the ${\alpha}$-factor preprosequence and a classical mammalian signal peptide derived from bovine ${\beta}$-casein was compared. The results showed that although the bovine ${\beta}$-casein signal peptide could direct high-level secretion of recombinant xylanase, it was relatively less efficient than the ${\alpha}$-factor preprosequence. In contrast, the bovine ${\beta}$-casein signal peptide caused remarkably more recombinant xylanase trapped intracellularly. Real-time RT-PCR analysis indicated that the difference in the secretory level between the two signal sequences was not due to the difference in the transcriptional efficiency.

An Analysis of Emergency Care Based on Prehospital Care Reports (일부 구급대의 응급처치활동 분석 - 구급활동일지를 중심으로 -)

  • Uhm, Tai-Hwan
    • The Korean Journal of Emergency Medical Services
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    • v.9 no.1
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    • pp.101-109
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    • 2005
  • The purpose of this study which was done by 250 Prehospital Care Reports(PCRs) survey of some squads in Seoul Metropolitan Fire & Disaster Management Department was to improve prehospital emergency care by means of quality management. The data were collected in 3 squads from Jun. 21 to Jul. 18, 2004 and analyzed by using SPSS Win 12.0 Version. The conclusions from this study were summarized as follows. The mean time of Event to treatment interval was $4.6{\pm}4.3$ minutes and 49.2% arrived at patient within 4 minutes. Platinum minute was observed 61.1% of verbal response, 73.3% of painful response, 77.8% of unresponsive. The great majority of patients couldn't receive advanced life support on account of limited scope of practice and strict direct medical control in the Emergency Medical Services Act. Data from quality improvement activity will be useful to expand indirect medical control which is able to activate prehospital care. To utilize PCR for quality improvement. It has to have data elements, run data, patient data, check boxes, narrative including US DOT's minimum data set.

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Genetic Identification of Spirometra erinaceieuropaei Spargana in Liaoning and Hubei Provinces, PR China

  • He, Li;Fang, Zheng-Ming;Xue, Ting;Zhang, Er-Fu;An, Chun-Li
    • Parasites, Hosts and Diseases
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    • v.57 no.3
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    • pp.309-312
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    • 2019
  • Spargana were collected from human and frogs in Liaoning and Hubei Provinces, China. PCR amplification and direct sequencing of A cox1 fragment was PCR-amplified from genomic DNA extracted from 7 specimens (5 from humans and 2 from frogs). The cox1 fragment (390 bp) showed 97-100% similarity to the reference sequence of S. erinaceieuropaei and 88-89% to the reference sequence of S. decipiens. There were 1-12 bases different between these worms, but no obvious genetic variation (0-3.3%) to the references. There was little difference of cox1 gene between sparganum samples of humans and frogs (1-3%). This study is the first report on S. erinaceieuropaei spargana from humans in Liaoning and Hubei Provinces.

Direct Stem Blot Immunoassay (DSBIA): A Rapid, Reliable and Economical Detection Technique Suitable for Testing Large Number of Barley Materials for Field Monitoring and Resistance Screening to Barley mild mosaic virus and Barley yellow mosaic virus

  • Jonson, Gilda;Park, Jong-Chul;Kim, Yang-Kil;Kim, Mi-Jung;Lee, Mi-Ja;Hyun, Jong-Nae;Kim, Jung-Gon
    • The Plant Pathology Journal
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    • v.23 no.4
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    • pp.260-265
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    • 2007
  • Testing a large number of samples from field monitoring and routine indexing is cumbersome and the available virus detection tools were labor intensive and expensive. To circumvent these problems we established tissue blot immunoassay (TBIA) method an alternative detection tool to detect Barley mild mosaic virus (BaMMV) and Barley yellow mosaic virus (BaYMV) infection in the field and greenhouse inoculated plants for monitoring and routine indexing applications, respectively. Initially, leaf and stem were tested to determine suitable plant tissue for direct blotting on nitrocellulose membrane. The dilutions of antibodies were optimized for more efficient and economical purposes. Results showed that stem tissue was more suitable for direct blotting for it had no background that interferes in the reaction. Therefore, this technique was referred as direct stem blot immunoassay or DSBIA, in this study. Re-used diluted (1:1000) antiserum and conjugate up to 3 times with the addition of half strength amount of concentrated antibodies was more effective in detecting the virus. The virus blotted on the nitrocellulose membrane from stem tissues kept at room temperature for 3 days were still detectable. The efficiency of DSBIA and RT-PCR in detecting BaMMV and BaYMV were relatively comparable. Results further proved that DSBIA is a rapid, reliable and economical detection method suitable for monitoring BaMMV and BaYMV infection in the field and practical method in indexing large scale of barley materials for virus resistance screening.

Evaluation of Yeast Diversity During Wine Fermentations with Direct Inoculation and pied de cuve Method at an Industrial Scale

  • Li, Erhu;Liu, Chuanhe;Liu, Yanlin
    • Journal of Microbiology and Biotechnology
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    • v.22 no.7
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    • pp.960-966
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    • 2012
  • The diversity and composition of yeast populations may greatly impact wine quality. This study investigated the yeast microbiota in two different types of wine fermentations: direct inoculation of a commercial starter versus pied de cuve method at an industrial scale. The pied de cuve fermentation entailed growth of the commercial inoculum used in the direct inoculation fermentation for further inoculation of additional fermentations. Yeast isolates were collected from different stages of wine fermentation and identified to the species level using Wallersterin Laboratory nutrient (WLN) agar followed by analysis of the 26S rDNA D1/D2 domain. Genetic characteristics of the Saccharomyces cerevisiae strains were assessed by a rapid PCR-based method, relying on the amplification of interdelta sequences. A total of 412 yeast colonies were obtained from all fermentations and eight different WL morphotypes were observed. Non-Saccharomyces yeast mainly appeared in the grape must and at the early stages of wine fermentation. S. cerevisiae was the dominant yeast species using both fermentation techniques. Seven distinguishing interdelta sequence patterns were found among S. cerevisiae strains, and the inoculated commercial starter, AWRI 796, dominated all stages in both direct inoculation and pied de cuve fermentations. This study revealed that S. cerevisiae was the dominant species and an inoculated starter could dominate fermentations with the pied de cuve method under controlled conditions.

Application of the 18S Ribosomal DNA (rDNA) PCR-RFLP Technique for the Differential Diagnosis of Anisakidosis (고래회충유충증 감별 진단을 위한 18S ribosomal DNA (rDNA) PCR-RFLP 법 적용)

  • Kim, Sun-Mee;Cho, Min-Kyung;Yu, Hak-Sun;Cha, Hee-Jae;Ock, Mee-Sun
    • Journal of Life Science
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    • v.19 no.9
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    • pp.1328-1332
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    • 2009
  • Anisakidosis is caused by anisakid nematodes (family Anisakidae) larvae which can cause not only direct tissue damage but also a severe allergic response related to excretory-secretion products. Lots of different species of anisakid larvae, including Anisakis simplex, Contracaecum, Goezia, Pseudoterranova, and Hysterothylacium, cause the anisakidosis. But it is difficult to diagnosis the species of larvae since the morphologies of larval anisakid nematodes are almost indistinguishable. In order to diagnosis the differential infections of larval anisakid nematodes, polymerase chain reaction - restriction fragment length polymorphism (PCR-RFLP) of 18S rDNA - was conducted. Three major species of anisakid larvae including A. simplex, C.ontracaecum spp, and Goezia spp. were collected from mackerel (Scomber japonicus), mullet (Mugil cephalus), founder (Paralichthys olivaceus), eel (Astroconger myriaster) and red sea bream (Pagrus major). PCR amplified 18S rDNA from each species of anisakid larvae was digested with eight restriction enzymes including Taq I, Hinf I, Hha I, Alu I, Dde I, Hae III, Sau96 I, and Sau3A I. The original sizes of PCR amplified 18S rDNA were 2.0Kb in both anisakid larvaes and Goezia. Restrction enzymes including Hinf 1, Alu 1, Hha I, Dde 1 and Hae III cut differently and distinguished the A. simplex and Contracaecum type C'. However, Contracaecum type A showed two different restriction enzyme cutting patterns by Taq 1, Hinf I, Alu 1, and Dde 1. One of the patterns was the same as those of A. simplex, Contracaecum type C' and Goezia and the other was unique. These results suggest that PCR-RFLP pattern by Hinf 1, Alu 1, Hae I, Dde 1 and Hae III can be applied to differential diagnosis of human infection with A. simplex and Contracaecum type C'. Contracaecum type A needs further study of classification by morphological characteristics and genetic analysis.

Mutation Analysis of Korean Patients with Glycogen Storage Disease Type Ia (한국인 당원병 제 Ia형 환자의 돌연변이 분석)

  • Kim, Jong-Won;Park, Ji-Yeon;Seo, Jeong-Kee
    • Pediatric Gastroenterology, Hepatology & Nutrition
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    • v.4 no.2
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    • pp.213-217
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    • 2001
  • Purpose: Glycogen storage disease type Ia (GSD Ia) is an autosomal recessive disorder of glycogen metabolism caused by glucose-6-phosphatase (G6Pase) deficiency. The clinical manifestations of G6Pase deficiency include growth retardation, hepatomegaly, hypoglycemia, lactic acidemia, hyperlipidemia and hyperuricemia. Many mutations of this gene have been found worldwide in various ethnic groups, establishing the molecular basis of GSD Ia. To elucidate a spectrum of the G6Pase gene mutations in Korean, we analyzed mutations in Korean patients with GSD Ia. Methods: Both alleles of 9 unrelated GSD 1a patients were studied by PCR and direct DNA sequencing methods. In all patients, GSD 1a was diagnosed by the enzyme assay for the liver biopsy specimen. Results: In Korean, the most prevalent mutation was g727t substitution in exon 5, which has been reported to cause abnormal mRNA splicing: Sixteen out of 18 alleles were found to have this mutation. In addition, we identified one novel mutation, a c611g, converting a proline to an alanine at codon 178. Conclusion: Our findings suggest that a screening for the g727t mutation by noninvasive molecular method can detect most cases of GSD Ia in Korean patients.

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Association between a p73 Gene Polymorphism and Genetic Susceptibility to Non-small Cell Lung Cancer in the South of China

  • Wang, Shuang-Shuang;Guo, Hai-Yan;Dong, Lin-Li;Zhu, Xiang-Qian;Ma, Liang;Li, Wen;Tang, Jian-Xin
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.23
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    • pp.10387-10391
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    • 2015
  • Background: This study aimed to identify any association between the p73 gene G4C14-to-A4T14 polymorphism and risk of non-small cell lung cancer (NSCLC) in the south of China. Materials and Methods: We genotyped the p73 gene polymorphism of peripheral blood DNA from 168 patients with NSCLC and 195 normal controls using HRM (high resolution melting) and PCR-CTPP (polymerase chain reaction with confronting two-pair primers). Results: The results of genotyping by HRM and PCR-CTPP were consistent with direct sequencing, the p73 genotype distribution in 168 lung cancer patients being as follows: GC/GC 101 cases (60.1%), GC/AT 59 cases (35.1%), AT/AT 8 cases (4.8%). The carriers of AT/AT genotype had a significantly reduced risk of NSCLC (OR=0.370; 95%CI: 0.170-0.806; p=0.010) as compared with non-carriers. However, we found no relations between p73 genotypes and histological type (p=0.798, $x^2=0.452$), tumor stage (p=0.806, $x^2=0.806$), or lymph node metastasis (p=0.578, $x^2=1.098$). Conclusions: Our findings suggest that the p73 G4C14-to-A4T14 polymorphism may be a modifier of NSCLC susceptibility in the Chinese population.