scholarly journals Detection and quantification of five major periodontal pathogens by single copy gene-based real-time PCR

2009 ◽  
Vol 15 (4) ◽  
pp. 195-204 ◽  
Author(s):  
Kati Hyvärinen ◽  
Saara Laitinen ◽  
Susanna Paju ◽  
Anne Hakala ◽  
Liisa Suominen-Taipale ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Single Copy ◽  
Single Copy Gene ◽  
Periodontal Pathogens ◽  
Copy Gene ◽  
Detection And Quantification

scholarly journals Real-Time PCR for Molecular Detection of Zoonotic and Non-Zoonotic Giardia spp. in Wild Rodents

2021 ◽  
Vol 9 (8) ◽  
pp. 1610
Author(s):  
Christian Klotz ◽  
Elke Radam ◽  
Sebastian Rausch ◽  
Petra Gosten-Heinrich ◽  
Toni Aebischer
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Gastrointestinal Disease ◽  
Single Copy ◽  
Marker Genes ◽  
Small Rodent ◽  
Analytical Sensitivity ◽  
Single Copy Gene ◽  
Wild Rodents ◽  
Copy Gene

Giardiasis in humans is a gastrointestinal disease transmitted by the potentially zoonotic Giardia duodenalis genotypes (assemblages) A and B. Small wild rodents such as mice and voles are discussed as potential reservoirs for G. duodenalis but are predominantly populated by the two rodent species Giardia microti and Giardia muris. Currently, the detection of zoonotic and non-zoonotic Giardia species and genotypes in these animals relies on cumbersome PCR and sequencing approaches of genetic marker genes. This hampers the risk assessment of potential zoonotic Giardia transmissions by these animals. Here, we provide a workflow based on newly developed real-time PCR schemes targeting the small ribosomal RNA multi-copy gene locus to distinguish G. muris, G. microti and G. duodenalis infections. For the identification of potentially zoonotic G. duodenalis assemblage types A and B, an established protocol targeting the single-copy gene 4E1-HP was used. The assays were specific for the distinct Giardia species or genotypes and revealed an analytical sensitivity of approximately one or below genome equivalent for the multi-copy gene and of about 10 genome equivalents for the single-copy gene. Retesting a biobank of small rodent samples confirmed the specificity. It further identified the underlying Giardia species in four out of 11 samples that could not be typed before by PCR and sequencing. The newly developed workflow has the potential to facilitate the detection of potentially zoonotic and non-zoonotic Giardia species in wild rodents.

The utility and application of real-time PCR and FISH in the detection of single-copy gene targets inEscherichia coliO157:H7 andSalmonellaTyphimurium

2010 ◽  
Vol 56 (3) ◽  
pp. 254-262 ◽  
Author(s):  
Merriam Haffar ◽  
Kimberley A. Gilbride
Keyword(s):  
Real Time ◽  
Dna Sequences ◽  
Real Time Pcr ◽  
Receptor Gene ◽  
Detection Limits ◽  
Single Copy ◽  
Target Cells ◽  
Single Copy Gene ◽  
Gene Copies ◽  
Copy Gene

The ultimate specificity in molecular-based assays for pathogen detection relies on the design of the primers and probes. Their ability to hybridize to DNA sequences found only in pathogens can be realized by designing primers and probes that are complementary to pathogen-specific virulence genes. This study evaluates the detection and enumeration strengths of real-time PCR (qPCR) and fluorescent in situ hybridization (FISH) for selected waterborne pathogens and their ultimate applicability within a monitoring framework. Detection limits calculated in the qPCR assay were 150 tir (intimin protein receptor) gene copies for Escherichia coli O157:H7 and 2 × 103invA (inner membrane invasive protein) gene copies for Salmonella enterica serovar Typhimurium. Detection limits were, however, at least 100-fold less sensitive in wastewater extracts, partly because of the inhibitory effect of the wastewater itself. Fluorescent signals from hybridized whole target cells were below the detection limit of the FISH assay. While this research demonstrates the potential detection strength of qPCR, it highlights the need for strong dependable primer and probe sets among PCR and FISH methodologies as well as the need for further signal amplification with DNA-targeted FISH for single-copy gene targets within environmental samples.

scholarly journals Quantitative real-time polymerase chain reaction based on single copy gene sequence for detection of periodontal pathogens

2004 ◽  
Vol 31 (12) ◽  
pp. 1054-1060 ◽  
Author(s):  
Juan Manuel Morillo ◽  
Laura Lau ◽  
Mariano Sanz ◽  
David Herrera ◽  
Conchita Martin ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
Gene Sequence ◽  
Single Copy ◽  
Single Copy Gene ◽  
Periodontal Pathogens ◽  
Chain Reaction ◽  
Copy Gene ◽  
Polymerase Chain

Detection and quantification of Histomonas meleagridis by real-time PCR targeting single copy genes

2015 ◽  
Vol 212 (3-4) ◽  
pp. 382-388 ◽  
Author(s):  
Imtiaz Hussain ◽  
Barbara Jaskulska ◽  
Michael Hess ◽  
Ivana Bilic
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Single Copy ◽  
Pcr Targeting ◽  
Detection And Quantification

Identification of a single-copy gene encoding a Type I chlorophyll a/b-binding polypeptide of photosystem I in Arabidopsis thaliana

1992 ◽  
Vol 84 (4) ◽  
pp. 561-567 ◽  
Author(s):  
Poul E. Jensen ◽  
Michael Kristensen ◽  
Tine Hoff ◽  
Jan Lehmbeck ◽  
Bjarne M. Stummann ◽  
...  
Keyword(s):  
Arabidopsis Thaliana ◽  
Chlorophyll A ◽  
Photosystem I ◽  
Single Copy ◽  
Type I ◽  
Single Copy Gene ◽  
Gene Encoding ◽  
Copy Gene

A Mouse Single-Copy Gene,Gtf2i,the Homolog of HumanGTF2I,That Is Duplicated in the Williams–Beuren Syndrome Deletion Region

Genomics ◽  
1998 ◽  
Vol 48 (2) ◽  
pp. 163-170 ◽  
Author(s):  
Yu-Ker Wang ◽  
Luis A. Pérez-Jurado ◽  
Uta Francke
Keyword(s):  
Single Copy ◽  
Single Copy Gene ◽  
Deletion Region ◽  
Copy Gene

scholarly journals Single-copy detection of somatic variants from solid and liquid biopsy

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Ana-Luisa Silva ◽  
Paulina Klaudyna Powalowska ◽  
Magdalena Stolarek ◽  
Eleanor Ruth Gray ◽  
Rebecca Natalie Palmer ◽  
...  
Keyword(s):  
Real Time ◽  
Single Molecule ◽  
Real Time Pcr ◽  
Clinical Decision Making ◽  
High Sensitivity ◽  
Clinical Decision ◽  
Single Copy ◽  
Turnaround Time ◽  
Copy Detection ◽  
Allele Specific

AbstractAccurate detection of somatic variants, against a background of wild-type molecules, is essential for clinical decision making in oncology. Existing approaches, such as allele-specific real-time PCR, are typically limited to a single target gene and lack sensitivity. Alternatively, next-generation sequencing methods suffer from slow turnaround time, high costs, and are complex to implement, typically limiting them to single-site use. Here, we report a method, which we term Allele-Specific PYrophosphorolysis Reaction (ASPYRE), for high sensitivity detection of panels of somatic variants. ASPYRE has a simple workflow and is compatible with standard molecular biology reagents and real-time PCR instruments. We show that ASPYRE has single molecule sensitivity and is tolerant of DNA extracted from plasma and formalin fixed paraffin embedded (FFPE) samples. We also demonstrate two multiplex panels, including one for detection of 47 EGFR variants. ASPYRE presents an effective and accessible method that simplifies highly sensitive and multiplexed detection of somatic variants.

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