A Fast Real-Time Polymerase Chain Reaction Method for Sensitive and Specific Detection of the Neisseria gonorrhoeae porA Pseudogene Stig Ove Hjelmevoll,

Slides:

Advertisements

Similar presentations

Measurement of Relative Copy Number of CDKN2A/ARF and CDKN2B in Bladder Cancer by Real-Time Quantitative PCR and Multiplex Ligation-Dependent Probe Amplification.

Advertisements

Development of a Novel One-Tube Isothermal Reverse Transcription Thermophilic Helicase-Dependent Amplification Platform for Rapid RNA Detection James Goldmeyer,

Clinical Laboratory Analysis of Immunoglobulin Heavy Chain Variable Region Genes for Chronic Lymphocytic Leukemia Prognosis Philippe Szankasi, David.

Quantitative Detection and Differentiation of Human Herpesvirus 6 Subtypes in Bone Marrow Transplant Patients by Using a Single Real-Time Polymerase Chain.

Clinical Validation of a New Triplex Real-Time Polymerase Chain Reaction Assay for the Detection and Discrimination of Herpes simplex Virus Types 1 and.

A Prospective Clinical Study on the Use of Reverse Transcription-Polymerase Chain Reaction for the Early Diagnosis of Dengue Fever Kamaljit Singh, Ajit.

Detection of Exon 12 Mutations in the JAK2 Gene

Direct DNA Amplification from Crude Clinical Samples Using a PCR Enhancer Cocktail and Novel Mutants of Taq Zhian Zhang, Milko B. Kermekchiev, Wayne.

The Use of COLD-PCR and High-Resolution Melting Analysis Improves the Limit of Detection of KRAS and BRAF Mutations in Colorectal Cancer Irene Mancini,

A Highly Sensitive, Multiplex Broad-Spectrum PCR-DNA-Enzyme Immunoassay and Reverse Hybridization Assay for Rapid Detection and Identification of Chlamydia.

Pre-Clinical Validation of a Novel, Highly Sensitive Assay to Detect PML-RARα mRNA Using Real-Time Reverse-Transcription Polymerase Chain Reaction James.

Allele-Specific Polymerase Chain Reaction for the Imatinib-Resistant KIT D816V and D816F Mutations in Mastocytosis and Acute Myelogenous Leukemia Christopher.

One-Step Ligation on RNA Amplification for the Detection of Point Mutations Lei Zhang, Jingjing Wang, Mia Coetzer, Stephanie Angione, Rami Kantor, Anubhav.

Locked Nucleic Acids Can Enhance the Analytical Performance of Quantitative Methylation-Specific Polymerase Chain Reaction Karen S. Gustafson The Journal.

Direct DNA Amplification from Crude Clinical Samples Using a PCR Enhancer Cocktail and Novel Mutants of Taq Zhian Zhang, Milko B. Kermekchiev, Wayne.

Multiplex Detection of Ehrlichia and Anaplasma Species Pathogens in Peripheral Blood by Real-Time Reverse Transcriptase-Polymerase Chain Reaction Kamesh.

Real-Time Polymerase Chain Reaction for Detecting Bacterial DNA Directly from Blood of Neonates Being Evaluated for Sepsis Jeanne A. Jordan, Mary Beth.

Whole Genome Amplification for Array Comparative Genomic Hybridization Using DNA Extracted from Formalin-Fixed, Paraffin-Embedded Histological Sections

MethySYBR, a Novel Quantitative PCR Assay for the Dual Analysis of DNA Methylation and CpG Methylation Density Pang-Kuo Lo, Hanano Watanabe, Pi-Chun.

High-Fidelity DNA Polymerase Enhances the Sensitivity of a Peptide Nucleic Acid Clamp PCR Assay for K-ras Mutations Bjørnar Gilje, Reino Heikkilä, Satu.

Comparison of BIOMED-2 Versus Laboratory-Developed Polymerase Chain Reaction Assays for Detecting T-Cell Receptor-γ Gene Rearrangements Keyur P. Patel,

Clinical Validation of a New Triplex Real-Time Polymerase Chain Reaction Assay for the Detection and Discrimination of Herpes simplex Virus Types 1 and.

Establishment and Study of Different Real-Time Polymerase Chain Reaction Assays for the Quantification of Cells with Deletions of Chromosome 7 Elia Mattarucchi,

Detection of Cytomegalovirus in Whole Blood Using Three Different Real-Time PCR Chemistries Erica Vincent, Zhengming Gu, Markus Morgenstern, Candace.

Diagnostic Testing for Pandemic Influenza in Singapore

Multi-Probe Real-Time PCR Identification of Common Mycobacterium Species in Blood Culture Broth Suporn Foongladda, Suporn Pholwat, Boonchuay Eampokalap,

Philippe Szankasi, Mohamed Jama, David W. Bahler

Limitations and Practical Procedure in BclII-Ig Heavy Chain Gene Rearrangement Real- Time Quantitative Polymerase Chain Reaction Barbara Dessars, Pierre.

Single Nucleotide Polymorphism-Based System Improves the Applicability of Quantitative PCR for Chimerism Monitoring Egle Gineikiene, Mindaugas Stoskus,

Association of Clinical Status of Follicular Lymphoma Patients after Autologous Stem Cell Transplant and Quantitative Assessment of Lymphoma in Blood.

The Effect of Primer-Template Mismatches on the Detection and Quantification of Nucleic Acids Using the 5′ Nuclease Assay Ralph Stadhouders, Suzan D.

Novel Molecular Method for Detection of Bovine-Specific Central Nervous System Tissues as Bovine Spongiform Encephalopathy Risk Material in Meat and Meat.

Detection of Central Nervous System Leukemia in Children with Acute Lymphoblastic Leukemia by Real-Time Polymerase Chain Reaction Sharon R. Pine, Changhong.

Protocol for the Use of Polymerase Chain Reaction in the Detection of Intraocular Large B-Cell Lymphoma in Ocular Samples Aires Lobo, Narciss Okhravi,

Single-Run, Parallel Detection of DNA from Three Pneumonia-Producing Bacteria by Real-Time Polymerase Chain Reaction Reinhard B. Raggam, Eva Leitner,

Specific Detection of Cytokeratin 20-Positive Cells in Blood of Colorectal and Breast Cancer Patients by a High Sensitivity Real-Time Reverse Transcriptase-Polymerase.

William L. Gerald, M.D., Ph.D, 1954–2008

Low Copy Number DNA Template Can Render Polymerase Chain Reaction Error Prone in a Sequence-Dependent Manner Mansour Akbari, Marianne Doré Hansen, Jostein.

A Consensus on Fungal Polymerase Chain Reaction Diagnosis?

Real-Time Polymerase Chain Reaction for Detecting Bacterial DNA Directly from Blood of Neonates Being Evaluated for Sepsis Jeanne A. Jordan, Mary Beth.

Improved Real-Time Multiplex Polymerase Chain Reaction Detection of Methylenetetrahydrofolate Reductase (MTHFR) 677C>T and 1298A>C Polymorphisms Using.

Rapid Detection of Clonal T-Cell Receptor-β Gene Rearrangements in T-Cell Lymphomas Using the LightCycler-Polymerase Chain Reaction with DNA Melting Curve.

Keeping Up With the Next Generation

Multiplexed Detection of Anthrax-Related Toxin Genes

Differential DNA Methylation as a Tool for Noninvasive Prenatal Diagnosis (NIPD) of X Chromosome Aneuploidies Floriana Della Ragione, Paola Mastrovito,

Clinical Laboratory Analysis of Immunoglobulin Heavy Chain Variable Region Genes for Chronic Lymphocytic Leukemia Prognosis Philippe Szankasi, David.

Detection of Exon 12 Mutations in the JAK2 Gene

Catherine E. Keegan, Anthony A. Killeen

Isothermal Strand-Displacement Polymerase Reaction for Visual Detection of the Southeast Asian–Type Deletion of α-Thalassemia Luxin Yu, Wei Wu, Puchang.

Benjamin P. Song, Surbhi Jain, Selena Y. Lin, Quan Chen, Timothy M

Development of a Quantitative Real-Time Polymerase Chain Reaction Assay for the Detection of the JAK2 V617F Mutation Elizabeth C. Wolstencroft, Katy.

Detection of Common Disease-Causing Mutations in Mitochondrial DNA (Mitochondrial Encephalomyopathy, Lactic Acidosis with Stroke-Like Episodes MTTL

Design and Evaluation of a Real-Time PCR Assay for Quantification of JAK2 V617F and Wild-Type JAK2 Transcript Levels in the Clinical Laboratory Jason.

Polymerase Chain Reaction Detection of Kaposi's Sarcoma-Associated Herpesvirus- Optimized Protocols and Their Application to Myeloma Langxing Pan, Laura.

Diagnosis of Human Congenital Cytomegalovirus Infection by Amplification of Viral DNA from Dried Blood Spots on Perinatal Cards Lori Scanga, Shu Chaing,

Rapid and Sensitive Real-Time Polymerase Chain Reaction Method for Detection and Quantification of 3243A>G Mitochondrial Point Mutation Rinki Singh,

Low Incidence of Minor BRAF V600 Mutation-Positive Subclones in Primary and Metastatic Melanoma Determined by Sensitive and Quantitative Real-Time PCR

Amplification Refractory Mutation System, a Highly Sensitive and Simple Polymerase Chain Reaction Assay, for the Detection of JAK2 V617F Mutation in Chronic.

Multiplex PCR Detection of GSTM1, GSTT1, and GSTP1 Gene Variants

Cecily P. Vaughn, Elaine Lyon, Wade S. Samowitz

A Pyrosequencing-Based Assay for the Rapid Detection of the 22q11

A Novel Method to Compensate for Different Amplification Efficiencies between Patient DNA Samples in Quantitative Real-Time PCR Jules Meijerink, Caroline.

Optimized Allele-Specific Real-Time PCR Assays for the Detection of Common Mutations in KRAS and BRAF Alois H. Lang, Heinz Drexel, Simone Geller-Rhomberg,

Karen Snow-Bailey, Ph.D., 1961–2006

Nucleotide sequence alignment of porA gene sequences from the examined English Neisseria gonorrhoeae porA mutants, compared to the porA sequences of previously.

A Consensus on Fungal Polymerase Chain Reaction Diagnosis?

A Prospective Clinical Study on the Use of Reverse Transcription-Polymerase Chain Reaction for the Early Diagnosis of Dengue Fever Kamaljit Singh, Ajit.

Validation of Roche LightCycler Epstein-Barr Virus Quantification Reagents in a Clinical Laboratory Setting Margaret L. Gulley, Hongxin Fan, Sandra H.

Improved Detection of the KIT D816V Mutation in Patients with Systemic Mastocytosis Using a Quantitative and Highly Sensitive Real-Time qPCR Assay Thomas.

Multiplex Ligation-Dependent Probe Amplification Identification of Whole Exon and Single Nucleotide Deletions in the CFTR Gene of Hispanic Individuals.

Presentation transcript:

A Fast Real-Time Polymerase Chain Reaction Method for Sensitive and Specific Detection of the Neisseria gonorrhoeae porA Pseudogene Stig Ove Hjelmevoll, Merethe Elise Olsen, Johanna U. Ericson Sollid, Håkon Haaheim, Magnus Unemo, Vegard Skogen The Journal of Molecular Diagnostics Volume 8, Issue 5, Pages 574-581 (November 2006) DOI: 10.2353/jmoldx.2006.060024 Copyright © 2006 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

Figure 1 Nucleotide sequence alignment of partial N. gonorrhoeae consensus porA pseudogene compiled from 87 sequences34 and the corresponding N. meningitidis sequence. The boxed sequences on the flanks are the forward and reverse primer location, respectively. The boxed sequence in the middle is the location of the probe. The MC58 sequence is the partial porA sequence of the previously published whole-genome sequenced N. meningitidis strain MC58.42 The Journal of Molecular Diagnostics 2006 8, 574-581DOI: (10.2353/jmoldx.2006.060024) Copyright © 2006 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

Figure 2 Dilution series of commercially purified N. gonorrhoeae DNA (Tebu-bio) and detection in the N. gonorrhoeae porA pseudogene real-time PCR developed in the present study. The six 10-fold dilutions showed good linearity. The Journal of Molecular Diagnostics 2006 8, 574-581DOI: (10.2353/jmoldx.2006.060024) Copyright © 2006 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

Figure 3 Standard curve based on the dilution series in Figure 2. The slope is the basis for the amplification efficacy (E) calculation. In a 100% efficient PCR, the DNA will double per cycle or use 3.333 cycles to increase DNA copies by 10-fold (23.33 ≈ 10). The Journal of Molecular Diagnostics 2006 8, 574-581DOI: (10.2353/jmoldx.2006.060024) Copyright © 2006 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

Figure 4 IAC amplification with low-level N. gonorrhoeae. The figure shows the internal amplification control amplified in dilutions of increasing concentrations of N. gonorrhoeae DNA. The amplification lines that do not cross the thick line are negative due to larger amounts of N. gonorrhoeae DNA. The Journal of Molecular Diagnostics 2006 8, 574-581DOI: (10.2353/jmoldx.2006.060024) Copyright © 2006 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

Figure 5 Dilution series of N. gonorrhoeae DNA using two parallels from master mixes with and without IAC. There was no significant difference in the amplification of N. gonorrhoeae DNA between the two master mixes as all four parallels are approximately equal. The mastermix without IAC shows a slightly higher ΔRn than the master mix with IAC. The Journal of Molecular Diagnostics 2006 8, 574-581DOI: (10.2353/jmoldx.2006.060024) Copyright © 2006 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions