scholarly journals Development of a reverse transcription (RT) polymerase chain reaction (PCR) method for the detection of human norovirus in bivalve molluscs

2021 ◽  
Author(s):  
Manjusha Lekshmi ◽  
Sanath H. Kumar ◽  
Kooloth Valappil Rajendran ◽  
Binaya Bhusan Nayak
Keyword(s):  
Polymerase Chain Reaction ◽  
Reverse Transcription ◽  
Rapid Screening ◽  
Bivalve Molluscs ◽  
Chain Reaction ◽  
Reverse Transcription Pcr ◽  
Pcr Method ◽  
Polymerase Chain ◽  
Rna Polymerase Gene ◽  
Single Set

Abstract Noroviruses are significant seafood-borne pathogens, commonly associated with the consumption of filter feeding bivalve molluscs. Here, we report the development of a reverse transcription polymerase chain reaction (RT-PCR) method using primers based on the RNA-dependent RNA polymerase gene of norovirus genogroup II (NoV GII). Samples of bivalves were processed for the concentration of virus and extraction of RNA, followed by reverse transcription PCR. A total of 50 molluscan shellfish samples were analyzed, of which 16 samples yielded positive amplifications of norovirus nucleic acid. The PCR method described here, involving a single set of primers, is useful for rapid screening of shellfish for NoV GII.

Multiplex reverse transcription-polymerase chain reaction combined with on-chip electrophoresis as a rapid screening tool for candidate gene sets

2005 ◽  
Vol 26 (9) ◽  
pp. 1687-1691 ◽  
Author(s):  
Rainer Wittig ◽  
Rüdiger Salowsky ◽  
Stephanie Blaich ◽  
Stefan Lyer ◽  
Juehn S. Maa ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Candidate Gene ◽  
Reverse Transcription ◽  
Screening Tool ◽  
Rapid Screening ◽  
Chain Reaction ◽  
Gene Sets ◽  
Polymerase Chain ◽  
On Chip

scholarly journals Precipitation Solvents for RNA Extraction of Dengue Virus Type 3: Dimethylformamide, Ethylenediamintetraacetic Acid, and Ultrapure H2O

2019 ◽  
Vol 3 (2) ◽  
pp. 78
Author(s):  
Rizqidhana Juliana Putri ◽  
Teguh Hari Sucipto ◽  
Harsasi Setyawati ◽  
Siti Churrotin ◽  
Ilham Harlan Amarullah ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Dengue Virus ◽  
Dengue Fever ◽  
Reverse Transcription ◽  
Rna Extraction ◽  
Chain Reaction ◽  
Dengue Virus Type 3 ◽  
Pcr Method ◽  
Polymerase Chain ◽  
Type 3

Dengue is a disease caused by a virus from the family Flaviviradae, carried by a female mosquito of Aedes aegypti species. Dengue fever is widespread in the tropic areas. It caused by rainfall, temperature and unplanned urbanization. According to the ministry of health , almost all provinces in Indonesia are endemic areas of dengue fever. In 2014, up to mid-December Dengue Hemorrhagic Fever (DHF) patients in 34 provinces in Indonesia are 71,668 people and 641. This figure is lower than the previous year, 2013 with 112,511 people and 871 deaths . This disease consists of four types of serotypes, namely DENV-1, DENV-2, DENV-3, and DENV-4. This disease can be identified using a variety of methods, one of the method is Reverse Transcription - Polymerase Chain Reaction (RT-PCR) method. This study aims to determine the ability of Dimethylformamide (DMF), Ethylenediamintetraacetic Acid (EDTA), and Ultrapure H2O as the substitute of  Ethanol for precipitation in RNA extraction process. The sample used in this research obtained from Surabaya. RNA extraction itself can be done by using a special kit for RNA extraction. In Reverse Transcription - Polymerase Chain Reaction method, first RNA is extracted and then transcribed back (Reverse Transcription) which then form cDNA that later will be amplified by using PCR method. In this study used specific primers for dengue virus type 3 (DENV-3). The results of this study show that DMF, EDTA, and Ultrapure H2O can be used as the substitute of Ethanol for precipitation on RNA extraction. The result is evidenced by the formation of viral DNA bands on gel electrophoresis results.

Reverse-Transcription Polymerase Chain Reaction Detection of the Enteroviruses

1999 ◽  
Vol 123 (12) ◽  
pp. 1161-1169 ◽  
Author(s):  
JoséR. Romero
Keyword(s):  
Polymerase Chain Reaction ◽  
Sensitivity And Specificity ◽  
Reverse Transcription ◽  
Health Care Cost ◽  
Cost Savings ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Reverse Transcription Pcr ◽  
Polymerase Chain ◽  
Pcr Conditions

Abstract Objective.—This review focuses on commercial and in-house–developed reverse-transcription polymerase chain reaction (RT-PCR) assays used for the detection of enteroviral infections. In addition to providing details on the performance of RT-PCR, its specificity, and sensitivity, the clinical utility of this diagnostic method with specific reference to its impact on hospitalization and cost savings is addressed. Data Sources.—MEDLINE was searched for reports relating to RT-PCR detection of the enteroviruses in adults and children. The search was restricted to studies reported in English language journals. Study Selection.—Reports documenting detailed information regarding the RT-PCR conditions, primers, sensitivity, specificity and, if relevant, clinical impact were selected for analysis. Data Extraction.—Details regarding method of extraction of the enteroviral genome, the primers used, RT-PCR conditions, and sensitivity and specificity of the assay were extracted from the literature. For reports detailing the use of RT-PCR in the clinical management of enteroviral infections in children, the reduction in duration of hospitalization and health care cost savings were recorded. Data Synthesis.—Reverse-transcription PCR can increase the yield of detection of enteroviruses from cerebrospinal fluid by a mean of approximately 20% over tissue culture. Reverse-transcription PCR of cerebrospinal fluid has been shown to exhibit sensitivity and specificity values of 86% to 100% and 92% to 100%, respectively. Reductions of 1 to 3 days of hospitalization per patient are predicted if RT-PCR is used to diagnose enteroviral meningitis in children. Conclusions.—Reverse-transcription PCR detection of enteroviral infections is an extremely rapid, sensitive, and specific diagnostic modality. Both commercial assays and assays developed in-house appear to be equivalent with regard to sensitivity and specificity. Reverse-transcription PCR diagnosis of enteroviral infections in children could reduce the length of hospitalization and result in significant health care cost savings.

scholarly journals METHOD FOR FMD VIRUS 146S COMPONENT CONCENTRATION DETERMINATION WITH REAL-TIME REVERSE TRANSCRIPTION – POLYMERASE CHAIN REACTION IN VACCINE RAW MATERIALS

2018 ◽  
pp. 31-35
Author(s):  
M. I. Doronin ◽  
A. M. Timina ◽  
D. A. Lozovoy ◽  
V. A. Starikov ◽  
D. V. Mikhalishin ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Regression Model ◽  
Real Time ◽  
Reverse Transcription ◽  
Raw Materials ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Pcr Method ◽  
Polymerase Chain

Method for determination of foot-and-mouth disease (FMD) virus 146S component concentration with real-time reverse transcription – polymerase chain reaction (rtRT-PCR) in vaccine raw materials is developed. Negative regression model of  С146S = (30.269 – Ct)/4.155 type allowing determination of FMDV 146S particle concentrations based on the amplification threshold cycle values (Ct) is proposed. It has been experimentally proven that the quantity of the 146S component determined by the real-time RT-PCR method using developed regression model and contained in the inoculation dose of FMD vaccine confers protection to the vaccinated animals against generalized FMD of A, O, Asia-1 types. rtRT-PCR method is highly sensitive and allows rapid and highly reliable estimation of  the 146S antigen concentration in FMD vaccine. The method for 146S particle quantity determination by real-time RT-PCR using the regression model is reliable and demonstrates high correlation (95.5–99.0%) with the complement fixation test results.

scholarly journals Semiquantitative reverse transcription-polymerase chain reaction to evaluate the expression patterns of genes involved in the oestrogen pathway

2000 ◽  
Vol 24 (3) ◽  
pp. 433-440 ◽  
Author(s):  
JM Rey ◽  
P Pujol ◽  
P Callier ◽  
V Cavailles ◽  
G Freiss ◽  
...  
Keyword(s):  
Gene Expression ◽  
Polymerase Chain Reaction ◽  
Reverse Transcription ◽  
Expression Patterns ◽  
Mrna Level ◽  
Screening Method ◽  
Housekeeping Gene ◽  
Rapid Screening ◽  
Chain Reaction ◽  
Polymerase Chain

The increasing number of factors to be taken into account in the oestrogen transcriptional process has created a need to develop a rapid screening method to evaluate their role in physiology and pathology. Molecular biology techniques enable gene expression studies at the mRNA level with small amounts of tissues. We therefore developed a semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) technique using fluorescent oligonucleotides to analyse simultaneously a large panel of interrelated genes involved in the oestrogen transcriptional pathway using a moderately expressed housekeeping gene, the hypoxanthine phosphoribosyltransferase gene (HPRT), as the reference gene. Expression levels of oestrogen receptors (ERalpha, ERbeta), cofactors AIB1, RIP140, SMRT and the Fas-associated protein-tyrosine phophatase-1 (FAP-1) genes were evaluated in breast, endometrial and ovarian cancer cell lines and in three ERalpha-positive and three ERalpha-negative breast cancer tumours. This technique provides a rapid and reliable way to quantitate simultaneously numerous mRNAs of genes involved in the oestrogen pathway from small amounts of tissues.

scholarly journals Sensitive Method for Testing Peanut Seed Lots for Peanut stripe and Peanut mottle viruses by Immunocapture-Reverse Transcription-Polymerase Chain Reaction

Plant Disease ◽  
2000 ◽  
Vol 84 (5) ◽  
pp. 559-561 ◽  
Author(s):  
A. G. Gillaspie ◽  
R. N. Pittman ◽  
D. L. Pinnow ◽  
B. G. Cassidy
Keyword(s):  
Polymerase Chain Reaction ◽  
Reverse Transcription ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Capsid Region ◽  
Large Numbers ◽  
Peanut Stripe Virus ◽  
Pcr Method ◽  
Polymerase Chain

An immunocapture-reverse transcription-polymerase chain reaction (IC-RT-PCR) method was developed for testing peanut (Arachis hypogaea) seed lots for infection by Peanut stripe virus (PStV) and Peanut mottle virus (PeMV). A small slice was removed from each seed distal to the radicle of a random 100-seed sample, the slices were extracted in buffer and centrifuged, and a portion of the supernatant was incubated in a tube that had been coated with antiserum to either PStV or PeMV. Following immunocapture of the virus, the tube was washed, the RT-PCR mix (with primers designed from conserved sequences within the capsid region of each virus) was placed in the same tubes, and the test completed. Results obtained on 15 previously untested seed lots from the collection indicated good correlation between virus detected by the IC-RT-PCR method and virus detected from the same seed lots by enzyme-linked immunosorbent assay (ELISA). The IC-RT-PCR method detected three lots infected with PeMV and none with PStV from 106 seed lots grown in Ecuador (results confirmed by ELISA). The IC-RT-PCR method is more sensitive than ELISA (currently used on samples consisting of five seeds), is useful for testing large numbers of seed lots of peanut germ plasm, and could be adapted to test other plants and detect other viruses.

scholarly journals METHOD FOR FMD VIRUS 146S COMPONENT CONCENTRATION DETERMINATION WITH REAL-TIME REVERSE TRANSCRIPTION – POLYMERASE CHAIN REACTION IN VACCINE RAW MATERIALS

2018 ◽  
pp. 26-30 ◽  
Author(s):  
M. I. Doronin ◽  
A. M. Timina ◽  
D. A. Lozovoy ◽  
V. A. Starikov ◽  
D. V. Mikhalishin ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Regression Model ◽  
Real Time ◽  
Reverse Transcription ◽  
Raw Materials ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Pcr Method ◽  
Polymerase Chain

Method for determination of foot-and-mouth disease (FMD) virus 146S component concentration with real-time reverse transcription – polymerase chain reaction (rtRT-PCR) in vaccine raw materials is developed. Negative regression model of  С146S = (30.269 – Ct )/4.155 type allowing determination of FMDV 146S particle concentrations based on the amplification threshold cycle values (Ct ) is proposed. It has been experimentally proven that the quantity of the 146S component determined by the real-time RT-PCR method using developed regression model and contained in the inoculation dose of FMD vaccine confers protection to the vaccinated animals against generalized FMD of A, O, Asia-1 types. rtRT-PCR method is highly sensitive and allows rapid and highly reliable estimation of the 146S antigen concentration in FMD vaccine. The method for 146S particle quantity determination by real-time RT-PCR using the regression model is reliable and demonstrates high correlation (95.5–99.0%) with the complement fixation test results.

1504: Molecular Diagnosis and Staging of Prostate Cancer Using Clusterin in Real Time Reverse Transcription Polymerase Chain Reaction (RT-PCR)

2006 ◽  
Vol 175 (4S) ◽  
pp. 485-486
Author(s):  
Sabarinath B. Nair ◽  
Christodoulos Pipinikas ◽  
Roger Kirby ◽  
Nick Carter ◽  
Christiane Fenske
Keyword(s):  
Prostate Cancer ◽  
Polymerase Chain Reaction ◽  
Real Time ◽  
Molecular Diagnosis ◽  
Reverse Transcription ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Polymerase Chain
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