A novel approach for the detection of DNA using immobilized peptide nucleic acid (PNA) probes and signal enhancement by real-time immuno-polymerase chain reaction (RT-iPCR)

2009 ◽  
Vol 394 (2) ◽  
pp. 529-537 ◽  
Author(s):  
Lillian Roth ◽  
Jutta Zagon ◽  
Anke Ehlers ◽  
Lothar W. Kroh ◽  
Hermann Broll
Keyword(s):  
Polymerase Chain Reaction ◽  
Nucleic Acid ◽  
Real Time ◽  
Peptide Nucleic Acid ◽  
Signal Enhancement ◽  
Chain Reaction ◽  
Novel Approach ◽  
Polymerase Chain ◽  
Immobilized Peptide

scholarly journals Simple and rapid detection of common fetal aneuploidies using peptide nucleic acid probe-based real-time polymerase chain reaction

2022 ◽  
Vol 12 (1) ◽  
Author(s):  
Subeen Hong ◽  
Seung Mi Lee ◽  
Sohee Oh ◽  
So Yeon Kim ◽  
Young Mi Jung ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Nucleic Acid ◽  
Real Time ◽  
Real Time Pcr ◽  
Peptide Nucleic Acid ◽  
Melting Curve ◽  
Peak Height ◽  
Chain Reaction ◽  
Pcr Assay ◽  
Polymerase Chain

AbstractTo examine the detection performance of a peptide nucleic acid (PNA) probe-based real-time time polymerase chain reaction (PCR) assay to detect common aneuploidies. Using amniotic fluid samples, PNA probe based real-time PCR (Patio DEP Detection Kit; SeaSun Biomaterials, Korea) assay was performed. PNA probe was designed to hybridize to similar sequences located on different segments of target chromosomes (21, 18, and 13) and a reference chromosome. Amplification of target sequences and melting curve analysis was performed. When analyzing the melting curve, the ratio of the peak height of the target and reference chromosome was calculated and determined as aneuploidy if the ratio of peak height was abnormal. All the results from the PNA probe-based real-time PCR and melting curve analyses were compared to those from conventional karyotyping. Forty-two cases with common aneuploidies (24 of trisomy 21, 12 of trisomy 18, and 6 of trisomy 13) and 131 cases with normal karyotype were analyzed. When comparing the karyotyping results, the sensitivity and specificity of the PNA probe-based real-time PCR assay were both 100%. The level of agreement was almost perfect (k = 1.00). PNA real-time PCR assay is a rapid and easy method for detecting common aneuploidies.

scholarly journals Peptide Nucleic Acid (PNA)-Enhanced Specificity of a Dual-Target Real-Time Quantitative Polymerase Chain Reaction (RT-qPCR) Assay for the Detection and Differentiation of SARS-CoV-2 from Related Viruses

Diagnostics ◽  
2020 ◽  
Vol 10 (10) ◽  
pp. 775
Author(s):  
Won-Suk Choi ◽  
Ju Hwan Jeong ◽  
Halcyon Dawn G. Nicolas ◽  
Sol Oh ◽  
Khristine Joy C. Antigua ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Nucleic Acid ◽  
Real Time ◽  
Peptide Nucleic Acid ◽  
Detection Method ◽  
Quantitative Polymerase Chain Reaction ◽  
Qpcr Assay ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Dual Target

The threat posed by coronaviruses to human health has necessitated the development of a highly specific and sensitive viral detection method that could differentiate between the currently circulating severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and other SARS-related coronaviruses (SARSr-CoVs). In this study, we developed a peptide nucleic acid (PNA)-based real-time quantitative polymerase chain reaction (RT-qPCR) assay targeting the N gene to efficiently discriminate SARS-CoV-2 from other SARSr-CoVs in human clinical samples. Without compromising the sensitivity, this method significantly enhanced the specificity of SARS-CoV-2 detection by 100-fold as compared to conventional RT-qPCR. In addition, we designed an RT-qPCR method for the sensitive and universal detection of ORF3ab-E genes of SARSr-CoV with a limit of detection (LOD) of 3.3 RNA copies per microliter. Thus, the developed assay serves as a confirmative dual-target detection method. Our PNA-mediated dual-target RT-qPCR assay can detect clinical SARS-CoV-2 samples in the range of 18.10–35.19 Ct values with an 82.6–100% detection rate. Furthermore, our assay showed no cross-reactions with other coronaviruses such as human coronaviruses (229E, NL63, and OC43) and Middle East respiratory syndrome coronavirus, influenza viruses (Type B, H1N1, H3N2, HPAI H5Nx, and H7N9), and other respiratory disease-causing viruses (MPV, RSV A, RSV B, PIV, AdV, and HRV). We, thus, developed a PNA-based RT-qPCR assay that differentiates emerging pathogens such as SARS-CoV-2 from closely related viruses such as SARSr-CoV and allows diagnosis of infections related to already identified or new coronavirus strains.

Ribonuclease H-cleavable and recombinase-quenching fluorescent probes for the real-time detection of strand invasion based amplification

2019 ◽  
Vol 11 (43) ◽  
pp. 5568-5576
Author(s):  
Sonja Elf ◽  
Kevin E. Eboigbodin
Keyword(s):  
Polymerase Chain Reaction ◽  
Nucleic Acid ◽  
Real Time ◽  
Fluorescent Probes ◽  
Nucleic Acid Amplification ◽  
Ribonuclease H ◽  
Chain Reaction ◽  
Amplification Method ◽  
Polymerase Chain ◽  
Strand Invasion

SIBA is an established nucleic acid amplification method that is used as an alternative to polymerase chain reaction (PCR).

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