Development and validation of a real-time quantitative PCR assay for rapid identification of Bacillus anthracis in environmental samples

2010 ◽  
Vol 88 (5) ◽  
pp. 1179-1192 ◽  
Author(s):  
Léonid M. Irenge ◽  
Jean-François Durant ◽  
Herbert Tomaso ◽  
Paola Pilo ◽  
Jaran S. Olsen ◽  
...  
Keyword(s):  
Bacillus Anthracis ◽  
Real Time ◽  
Quantitative Pcr ◽  
Environmental Samples ◽  
Rapid Identification ◽  
Pcr Assay ◽  
Real Time Quantitative Pcr ◽  
Development And Validation ◽  
Quantitative Pcr Assay

Development and validation of a real-time quantitative PCR assay to detect Xanthomonas axonopodis pv. allii from onion seed

2015 ◽  
Vol 114 ◽  
pp. 78-86 ◽  
Author(s):  
Isabelle Robène ◽  
Marion Perret ◽  
Emmanuel Jouen ◽  
Aline Escalon ◽  
Marie-Véronique Maillot ◽  
...  
Keyword(s):  
Real Time ◽  
Quantitative Pcr ◽  
Pcr Assay ◽  
Xanthomonas Axonopodis ◽  
Real Time Quantitative Pcr ◽  
Onion Seed ◽  
Development And Validation ◽  
Quantitative Pcr Assay

scholarly journals Real-Time Quantitative PCR Assay for Monitoring of Nervous Necrosis Virus Infection in Grouper Aquaculture

2011 ◽  
Vol 49 (3) ◽  
pp. 1090-1096 ◽  
Author(s):  
H.-C. Kuo ◽  
T.-Y. Wang ◽  
P.-P. Chen ◽  
Y.-M. Chen ◽  
H.-C. Chuang ◽  
...  
Keyword(s):  
Virus Infection ◽  
Real Time ◽  
Quantitative Pcr ◽  
Nervous Necrosis Virus ◽  
Necrosis Virus ◽  
Pcr Assay ◽  
Real Time Quantitative Pcr ◽  
Quantitative Pcr Assay

scholarly journals Determination of RhD Zygosity: Comparison of a Double Amplification Refractory Mutation System Approach and a Multiplex Real-Time Quantitative PCR Approach

2001 ◽  
Vol 47 (4) ◽  
pp. 667-672 ◽  
Author(s):  
Rossa W K Chiu ◽  
Michael F Murphy ◽  
Carrie Fidler ◽  
Benny C Y Zee ◽  
James S Wainscoat ◽  
...  
Keyword(s):  
Real Time ◽  
Quantitative Pcr ◽  
Gene Dosage ◽  
Amplification Refractory Mutation System ◽  
Pcr Assay ◽  
Real Time Quantitative Pcr ◽  
Mutation System ◽  
Allele Specific ◽  
Quantitative Pcr Assay

Abstract Background: Rh isoimmunization and hemolytic disease of the newborn still occur despite the availability of Rh immunoglobulin. For the prenatal investigation of sensitized RhD-negative pregnant women, determination of the zygosity of the RhD-positive father has important implications. The currently available molecular methods for RhD zygosity assessment, in general, are technically demanding and labor-intensive. Therefore, at present, rhesus genotype assessment is most commonly inferred from results of serological tests. The recent elucidation of the genetic structure of the prevalent RHD deletion in Caucasians, as well as the development of real-time PCR, allowed us to explore two new approaches for the molecular determination of RhD zygosity. Methods: Two methods for RhD zygosity determination were developed. The first was based on the double Amplification Refractory Mutation System (double ARMS). The second was based on multiplex real-time quantitative PCR. For the double ARMS assay, allele-specific primers were designed to directly amplify the most prevalent RHD deletion found in RhD-negative individuals in the Caucasian population. The multiplex real-time quantitative PCR assay, on the other hand, involved coamplification and quantification of RHD-specific sequences in relation to a reference gene, albumin, in a single PCR reaction. A ratio, ΔCt, based on the threshold cycle, was then determined and reflects the RHD gene dosage. Results: The allele-specific primers of the double ARMS assay reliably amplified the RHD-deleted allele and therefore accurately distinguished homozygous from heterozygous RhD-positive samples. The results were in complete concordance with serological testing. For the multiplex real-time quantitative PCR assay, the ΔCt values clearly segregated into two distinct populations according to the RHD gene dosage, with mean values of 1.70 (SD, 0.17) and 2.62 (SD, 0.29) for the homozygous and heterozygous samples, respectively (P <0.001, t-test). The results were in complete concordance with the results of serological testing as well as with the double ARMS assay. Conclusion: Double ARMS and real-time quantitative PCR are alternative robust assays for the determination of RhD zygosity.

Sign in / Sign up

Export Citation Format

Share Document