• Title/Summary/Keyword: PAN-PCR

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Novel pan-lineage VP1 specific degenerate primers for precise genetic characterization of serotype O foot and mouth disease virus circulating in India

  • Sagar Ashok Khulape;Jitendra Kumar Biswal;Chandrakanta Jana;Saravanan Subramaniam;Rabindra Prasad Singh
    • Journal of Veterinary Science
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    • v.24 no.3
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    • pp.40.1-40.6
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    • 2023
  • Analysis of the VP1 gene sequence of the foot and mouth disease virus (FMDV) is critical to understanding viral evolution and disease epidemiology. A standard set of primers have been used for the detection and sequence analysis of the VP1 gene of FMDV directly from suspected clinical samples with limited success. The study validated VP1-specific degenerate primer-based reverse transcription polymerase chain reaction (RT-PCR) for the qualitative detection and sequencing of serotype O FMDV lineages circulating in India. The novel degenerate primer-based RT-PCR amplifying the VP1 gene can circumvent the genetic heterogeneity observed in viruses after cell culture adaptation and facilitate precise viral gene sequence analysis from clinical samples.

Infected Left Atrial Myxoma Presenting Without Bacterial Growth on Blood Cultures: A Case Report

  • Shi A Kim;WonKyung Pyo;Sung-Ho Jung
    • Journal of Chest Surgery
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    • v.56 no.2
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    • pp.136-139
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    • 2023
  • Although cardiac myxoma is one of the most common types of benign cardiac tumors, infected cardiac myxoma is very infrequent. The diagnosis of infected cardiac myxoma may be challenging because the presenting symptoms are non-specific and established management guidelines are lacking. This report describes a 39-year-old woman with a 5-month history of uncontrolled fever, chills, and myalgia who was diagnosed with myxoma and underwent mass excision. Although blood and urine cultures were negative for growing bacteria, a pathologic examination showed that the excised mass was a left atrial myxoma, with pan-bacterial polymerase chain reaction (PCR) of the surgical specimen revealing Haemophilus parainfluenzae at 99.87%, resulting in a diagnosis of infected cardiac myxoma. Laboratory tests, such as PCR, may supplement culture results in the diagnosis of infected cardiac myxoma.

Expression and Clinical Significance of Osteopontin in Calcified Breast Tissue

  • Huan, Jin-Liang;Xing, Li;Qin, Xian-Ju;Gao, Zhi-Guang;Pan, Xiao-Feng;Zhao, Zhi-Dong
    • Asian Pacific Journal of Cancer Prevention
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    • v.13 no.10
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    • pp.5219-5223
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    • 2012
  • Osteopontin (OPN) is an integrin-binding protein, believed to be involved in a variety of physiological cellular functions. The physiology of OPN is best documented in the bone where this secreted adhesive glycoprotein appears to be involved in osteoblast differentiation and bone formation. In our study, we used semi-quantitative RT-PCR of osteopontin in calcification tissue of breast to detect breast cancer metastasis. The obtained data indicate that the expression of osteopontin is related to calcification tissue of breast, and possibly with the incidence of breast cancer. The expression strength of OPN by RT-PCR detection was related to the degree of malignancy of breast lesions, suggesting a close relationship between OPN and breast calcification tissue. The results revealed that expression of OPN mRNA is related to calcification of breast cancer tissue and to the development of breast cancer. Determination of OPN mRNA expression can be expected to be a guide to clinical therapy and prediction of the prognosis of breast cancer patients.

Development of multiplex polymerase chain reaction for the detection of vancomycin resistant genotypes and Enterococcus Sp.-specific genes (장구균의 vancomycin 내성 유전자와 종 특이유전자의 검출을 위한 Multiplex polymerase chain reaction 개발)

  • Cho, Yun-Sang;Lee, Hee-Soo;Kim, Jong-Man;Ahn, Jong-Sam;Ryu, Pan-Dong;Park, Yong-Ho;Yoo, Han-Sang;Lee, Mun-Han
    • Korean Journal of Veterinary Research
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    • v.43 no.1
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    • pp.103-112
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    • 2003
  • A multiplex PCR assay, which allows simultaneous detection of vancomycin resistant genotypes and Enterococcus species-specific genes, was developed. Vancomycin resistant enterococci (VRE) from chickens and humans could be detected for vanA, vanB, vanC-1, vanC-2, $ddl_{E.faecium}$ and $ddl_{E.faecalis}$ by multiplex PCR. Eight isolates of VRE from humans (n=11) had $ddl_{E.faecium}$ and vanA, and 3 isolates of the VRE had $ddl_{E.faecium}$ and vanB. One isolate of VRE from chickens (n=6) had $ddl_{E.faecium}$ and vanA, and 5 isolates of the VRE had only vanA. E. faecium, E. faecalis, E. gallinarum and E. casseliflavus were also confirmed for the species-specific gene by multiplex PCR. This multiplex PCR could detect E. faecium, E. faecalis, E. gallinarum, E. casseliflavus, vanA, vanB, vanC-1 and vanC-2, simultaneously. The PCR assay established in the present study can be an alternative to time-consuming biochemical tests and antibiotic susceptibility tests of Enterococcus spp.

miR-186 Regulates Glycolysis through Glut1 During the Formation of Cancer-associated Fibroblasts

  • Sun, Pan;Hu, Jun-Wei;Xiong, Wu-Jun;Mi, Jun
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.10
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    • pp.4245-4250
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    • 2014
  • Emerging evidence has suggested that glycolysis is enhanced in cancer-associated fibroblasts (CAF), and miR-186 is downregulated during the CAF formation. However, it is not clear whether miR-186 is involved in the regulation of glycolysis and what the role of miR-186 plays during the CAF formation. In this study, quantitative PCR analysises show miR-186 is downregulated during the CAF formation. Moreover, miR-186 targets the 3' UTR of Glut1, and its overexpression results in the degradation of Glut1 mRNA, which eventually reduces the level of Glut1 protein. On the other hand, knockdown of miR-186 increased the expression of Glut1. Both time course and dose response experiments also demonstrated that the protein and mRNA levels of Glut1 increase during CAF formation, according to Western blot and quantitative PCR analyses, respectively. Most importantly, besides the regulation on cell cycle progression, miR-186 regulates glucose uptake and lactate production which is mediated by Glut1. These observations suggest that miR-186 plays important roles in glycolysis regulation as well as cell cycle checkpoint activation.

Upregulation and Clinicopathological Significance of Long Non-coding NEAT1 RNA in NSCLC Tissues

  • Pan, Lin-Jiang;Zhong, Teng-Fei;Tang, Rui-Xue;Li, Ping;Dang, Yi-Wu;Huang, Su-Ning;Chen, Gang
    • Asian Pacific Journal of Cancer Prevention
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    • v.16 no.7
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    • pp.2851-2855
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    • 2015
  • Background: Recent reports have shown that nuclear enriched abundant transcript 1 (NEAT1), a long noncoding RNA (lncRNA), contributes to the precise control of gene expression and is related to several human malignancies. However, limited data are available on the expression and function of NEAT1 in lung cancer. The major objective of the current study was to profile the expression and clinicopathological significance of NEAT1 in non-small cell lung cancers (NSCLCs). Materials and Methods: NEAT1 expression in 125 NSCLC cases and paired adjacent non-cancer tissues was assessed by real-time quantitative reverse transcription-PCR (qRT-PCR). Relationships between NEAT1 and clinicopathological factors were also investigated. Results: The relative level of NEAT1 was $6.98{\pm}3.74$ in NSCLC tissues, significantly elevated as compared to that of the adjacent non-cancer lung tissues ($4.83{\pm}2.98$, p<0.001). The area under curve (AUC) of high expression of NEAT1 to diagnose NSCLC was 0.684 (95% CI: 0.619~0.750, p<0.001). NEAT1 expression was positively correlated with patient age (r=-2.007, p=0.047), lymphatic metastasis (r=-2.731, p=0.007), vascular invasion (r=-3.617, p=0.001) and clinical TNM stage (r=-4.134, p<0.001). Conclusions: This study indicates that NEAT1 might be associated with oncogenesis and progression in NSCLC, and suggests application in molecular targeted therapy.

Genetic Diversity of Schistosoma haematobium Eggs Isolated from Human Urine in Sudan

  • Quan, Juan-Hua;Choi, In-Wook;Ismail, Hassan Ahmed Hassan Ahmed;Mohamed, Abdoelohab Saed;Jeong, Hoo-Gn;Lee, Jin-Su;Hong, Sung-Tae;Yong, Tai-Soon;Cha, Guang-Ho;Lee, Young-Ha
    • Parasites, Hosts and Diseases
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    • v.53 no.3
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    • pp.271-277
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    • 2015
  • The genetic diversity of Schistosoma haematobium remains largely unstudied in comparison to that of Schistosoma mansoni. To characterize the extent of genetic diversity in S. haematobium among its definitive host (humans), we collected S. haematobium eggs from the urine of 73 infected schoolchildren at 5 primary schools in White Nile State, Sudan, and then performed a randomly amplified polymorphic DNA marker ITS2 by PCR-RFLP analysis. Among 73 S. haematobium egg-positive cases, 13 were selected based on the presence of the S. haematobium satellite markers A4 and B2 in their genomic DNA, and used for RFLP analysis. The 13 samples were subjected to an RFLP analysis of the S. haematobium ITS2 region; however, there was no variation in size among the fragments. Compared to the ITS2 sequences obtained for S. haematobium from Kenya, the nucleotide sequences of the ITS2 regions of S. haematobium from 4 areas in Sudan were consistent with those from Kenya (> 99%). In this study, we demonstrate for the first time that most of the S. haematobium population in Sudan consists of a pan-African S. haematobium genotype; however, we also report the discovery of Kenyan strain inflow into White Nile, Sudan.

Evaluation of the Accuracy of the $EasyTest^{TM}$ Malaria Pf/Pan Ag, a Rapid Diagnostic Test, in Uganda

  • Chong, Chom-Kyu;Cho, Pyo Yun;Na, Byoung-Kuk;Ahn, Seong Kyu;Kim, Jin Su;Lee, Jin-Soo;Lee, Sung-Keun;Han, Eun-Taek;Kim, Hak-Yong;Park, Yun-Kyu;Cha, Seok Ho;Kim, Tong-Soo
    • Parasites, Hosts and Diseases
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    • v.52 no.5
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    • pp.501-505
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    • 2014
  • In recent years, rapid diagnostic tests (RDTs) have been widely used for malaria detection, primarily because of their simple operation, fast results, and straightforward interpretation. The Asan $EasyTest^{TM}$ Malaria Pf/Pan Ag is one of the most commonly used malaria RDTs in several countries, including Korea and India. In this study, we tested the diagnostic performance of this RDT in Uganda to evaluate its usefulness for field diagnosis of malaria in this country. Microscopic and PCR analyses, and the Asan $EasyTest^{TM}$ Malaria Pf/Pan Ag rapid diagnostic test, were performed on blood samples from 185 individuals with suspected malaria in several villages in Uganda. Compared to the microscopic analysis, the sensitivity of the RDT to detect malaria infection was 95.8% and 83.3% for Plasmodium falciparum and non-P. falciparum, respectively. Although the diagnostic sensitivity of the RDT decreased when parasitemia was ${\leq}500\;parasites/{\mu}l$, it showed 96.8% sensitivity (98.4% for P. falciparum and 93.8% for non-P. falciparum) in blood samples with parasitemia ${\geq}100\;parasites/{\mu}l$. The specificity of the RDT was 97.3% for P. falciparum and 97.3% for non-P. falciparum. These results collectively suggest that the accuracy of the Asan $EasyTest^{TM}$ Malaria Pf/Pan Ag makes it an effective point-of-care diagnostic tool for malaria in Uganda.

Expression of Zonular Occludens-1 in Mouse Testis (생쥐 정소내 Zonular Occludens-1 발현)

  • Gye, Myung-Chan;Lee, Yang-Han;Kim, Chang-gyem;Kim, Moon-Kyoo;Lee, Hang
    • Development and Reproduction
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    • v.4 no.1
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    • pp.37-43
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    • 2000
  • Spatiotemporal expression of two isoforms of zonular occludens-1 (ZO-1), tight junctional protein, was examined in mouse testis. By RT-PCR, transcripts encoding two isoforms of ZO-1; ZO-1$\alpha$+ and ZO-1$\alpha$- were detected in testis. Two different forms of ZO-1 antigens with Mr.225 and 2001 KDa were detected in western blot of extract of neonatal to adult testis, coinciding with the result of RT-PCR. The relative amount of ZO-1 $\alpha$- versus 20-1 $\alpha$+ increased as the mice matured. In immunostaining using the pan antibody which detected both isoforms, ZO-1 was localized in the intercellular spaces in the Sertoli cell - Sertoli cell contacts in periphery of seminiferous tubule as well as Sertoli cell - germ cells contacts within the seminiferous tubule. The expression of ZO-1 was ubiquitous in both junctional area and cytoplasm of seminiferous tubule components. However, more intense signals were found in Sertoli cell junctional areas according to sexual maturation. The changes in the relative amount of both isoforms and spatial distribution of ZO-1 at the periphery of seminiferous tubule might be important for functional appearance of blood testis barrier and spermatogenesis.

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