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Multiplex polymerase chain reaction

About: Multiplex polymerase chain reaction is a research topic. Over the lifetime, 6409 publications have been published within this topic receiving 221244 citations.


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Journal ArticleDOI
TL;DR: A real-time PCR assay that simultaneously detects four bacterial agents that could be used in bioterrorism is described, specifically designed to test sterile body fluids.
Abstract: The advent of bioterrorism has highlighted the need for rapid, simple, and robust diagnostic assays to detect select agents. Mortality from select agents may be greatly reduced by prompt treatment (1); however, treatment may be delayed if diagnostic assays are outsourced to reference laboratories. Most bacterial species that would likely be used as bioterrorism agents infect the blood stream during the course of life-threatening disease. Furthermore, even “nonseptic” syndromes may produce hematogenous bacterial DNA that could be detected by a sensitive assay (2). This means that a rapid “molecular” version of a blood culture would fulfill many of the rapid diagnostic needs for biodefense. Bacteria can be detected in blood and other sterile body sites by the identification of species-specific DNA sequences in their 16S rRNA genes. These species-specific sequences are flanked by conserved sequences, permitting most rRNA targets to be amplified by PCR using a limited set of “universal” primers (3). Real-time PCR is well suited for sensitive and specific pathogen detection because it is performed in hermetically sealed wells, which greatly reduces the risk of cross-contamination, and it does not require post-PCR analysis (4). Real-time PCR assays have been developed for some select agents, most of which use fluorogenic 5′-nuclease (TaqMan) probes (5)(6)(7). However, TaqMan probes are difficult to use in multiplex PCR assays (8)(9). In contrast, molecular beacons are real-time PCR probes that are particularly amenable to multiplexing (10). They can be labeled with differently colored fluorophores (11), use a common nonfluorescent quenching moiety (9), and have thermodynamic properties that favor highly specific detection of nucleic acid sequences (12). Here we describe a real-time PCR assay that simultaneously detects four bacterial agents that could be used in bioterrorism. This assay is specifically designed to test sterile body fluids, where a …

59 citations

Journal ArticleDOI
TL;DR: The development of a multiplex real-time reverse-transcriptase polymerase chain reaction (RRT-PCR) assay using SYBR Green 1 chemistry for universal detection of avian influenza viruses and specific subtyping of H9N2 isolates based on melting temperatures (T(m)) discriminations is highlighted.

59 citations

Journal ArticleDOI
01 Jul 1990-Genomics
TL;DR: The polymerase chain reaction is used to amplify multiple DNA fragments originating from dispersed genomic segments that are flanked by Alu repeats to create "alumorphs", which could be useful for linkage mapping of the human genome.

59 citations

Journal ArticleDOI
TL;DR: A multiplex polymerase chain reaction (PCR) protocol to detect the presence of four genes linked to avian E. coli virulence, many of which reside on a large transmissible plasmid, suggest that this protocol might be useful in characterization and study of avianE.
Abstract: SUMMARY. Colibacillosis caused by Escherichia coli infections account for significant morbidity and mortality in the poultry industry. Yet, despite the importance of colibacillosis, much about the virulence mechanisms employed by avian E. coli remains unknown. In recent years several genes have been linked to avian E. coli virulence, many of which reside on a large transmissible plasmid. In the present study, a multiplex polymerase chain reaction (PCR) protocol to detect the presence of four of these genes is described. Such a protocol may supplement current diagnostic schemes and provide a rapid means of characterizing the E. coli causing disease in poultry. The targets of this procedure included iss, the increased serum survival gene; tsh, the temperature sensitive hemagglutinin gene; cvi, the ColV immunity gene; and iucC, a gene of the aerobactin operon. Organisms, known for their possession or lack of these genes, were used as a source of the template DNA to develop the multiplex PCR protocol...

59 citations

Journal ArticleDOI
TL;DR: The mPCR has the potential to be useful for routine molecular diagnosis and epidemiology and was sensitive and specific in detecting each target agent in composite cell cultures and clinical specimens.
Abstract: A multiplex PCR (mPCR) assay was developed and evaluated for its ability to simultaneously detect multiple viral infections of swine. Specific primers were designed for each of the following four DNA or RNA viruses: porcine circovirus type 2 (PCV2), porcine parvovirus (PPV), pseudorabies virus (PRV), and porcine reproductive and respiratory syndrome virus (PRRSV). Each target produced a specific amplicon with a size of 353 bp (PCV2), 271 bp (PPV), 194 bp (PRV), or 434 bp (PRRSV). The assay was sensitive and specific in detecting each target agent in composite cell cultures and clinical specimens. Results from mPCR were confirmed by PCR for individual viruses and by virus isolation. In conclusion, the mPCR has the potential to be useful for routine molecular diagnosis and epidemiology.

59 citations


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Performance
Metrics
No. of papers in the topic in previous years
YearPapers
2023275
2022448
2021172
2020176
2019221
2018220