A highly sensitive and specific real-time quantitative PCR for BRAF V600E/K mutation screening

Lung, Jrhau; Hung, Ming‐Szu; Lin, Yu‐Ching; Jiang, Yuan; Yu, Fang; Lu, Ming‐Shian; Hsieh, Ching‐Chuan; Wang, Chia‐Siu; Kuan, Feng‐Che; Lu, Chang‐Hsien; Chen, Ping‐Tsung; Lin, Chieh‐Mo; Chou, Yen-Li; Yang, Tsung‐Ming; Chen, Fen Fen; Lin, Paul Y.; Hsieh, Meng‐Jer; Tsai, Ying Huang

doi:10.1038/s41598-020-72809-7

Cited by 8 publications

(7 citation statements)

References 44 publications

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“…Similarly, a highly sensitive and specific RT-qPCR method has been developed for screening BRAF V600E/K mutation, which frequently occurs in lung cancers. The technique is useful for studying the incidence and clinicopathological features of BRAF V600E/K mutation in lung cancer patients [ 28 ].…”

Section: Applicationsmentioning

confidence: 99%

Real-Time Polymerase Chain Reaction: Current Techniques, Applications, and Role in COVID-19 Diagnosis

Artika

Dewi

Nainggolan

et al. 2022

Genes

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Successful detection of the first SARS-CoV-2 cases using the real-time polymerase chain reaction (real-time PCR) method reflects the power and usefulness of this technique. Real-time PCR is a variation of the PCR assay to allow monitoring of the PCR progress in actual time. PCR itself is a molecular process used to enzymatically synthesize copies in multiple amounts of a selected DNA region for various purposes. Real-time PCR is currently one of the most powerful molecular approaches and is widely used in biological sciences and medicine because it is quantitative, accurate, sensitive, and rapid. Current applications of real-time PCR include gene expression analysis, mutation detection, detection and quantification of pathogens, detection of genetically modified organisms, detection of allergens, monitoring of microbial degradation, species identification, and determination of parasite fitness. The technique has been used as a gold standard for COVID-19 diagnosis. Modifications of the standard real-time PCR methods have also been developed for particular applications. This review aims to provide an overview of the current applications of the real-time PCR technique, including its role in detecting emerging viruses such as SARS-CoV-2.

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Section: Applicationsmentioning

confidence: 99%

Real-Time Polymerase Chain Reaction: Current Techniques, Applications, and Role in COVID-19 Diagnosis

Artika

Dewi

Nainggolan

et al. 2022

Genes

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“…Finally, although the LOD of the present approach is higher than those reported for other methods (Table S-4), − it demonstrated to be adequate for the unequivocal identification of a SNP from a fingerprick sample. Moreover, our method, which can be performed in less than 50 min (DNA extraction, 30 s; amplification, 15 min; SA-Poly HRP, 20 min; detection, 5 min) and under 37 °C isothermal conditions, achieves a considerable reduction in assay time, complexity, and costs when compared with other SNP detection methods as summarized in Tables S-4, S-5, and S-6.…”

Section: Resultsmentioning

confidence: 55%

Solid-Phase Primer Elongation Using Biotinylated dNTPs for the Detection of a Single Nucleotide Polymorphism from a Fingerprick Blood Sample

2021

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Isothermal recombinase polymerase amplification-based solid-phase primer extension is used for the optical detection of a hypertrophic cardiomyopathy associated single nucleotide polymorphism (SNP) in a fingerprick blood sample. The assay exploits four thiolated primers which have the same sequences with the exception of the 3′terminal base. Target DNA containing the SNP site hybridizes to all four of the immobilized probes, with primer extension only taking place from the primer containing the terminal base that is complementary to the SNP under interrogation. Biotinylated deoxynucleotide triphosphates are used in the primer extension, allowing postextension addition of streptavidin-poly-horseradish peroxidase to bind to the incorporated biotinylated dNTPs. The signal generated following substrate addition can then be measured optically. The percentage of biotinylated dNTPs and the duration of primer extension is optimized and the system applied to the identification of a SNP in a fingerprick blood sample. A methodology of thermal lysis using a 1 in 5 dilution of the fingerprick blood sample prior to application of 95 °C for 30 s is used to extract genomic DNA, which is directly used as a template for solid-phase primer extension on microtiter plates, followed by optical detection. The SNP in the fingerprick sample was identified and its identity corroborated using ion torrent next generation sequencing. Ongoing work is focused on extension to the multiplexed detection of SNPs in fingerprick and other biological samples.

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“…Third, relationships among cells are lost in thin sections unless the cells of interest happen to fall into the same 5 micron plane. For example, the relationship between an immune cell and a neoplastic cell can not be appreciated unless the two cells happen to fall in the same plane of section (Lin et al, 2022). Fourth, the exact point at which an event occurs is not identifiable in 2D slides.…”

Section: Introductionmentioning

confidence: 99%

Tissue clearing and 3D reconstruction of digitized, serially sectioned slides provide novel insights into pancreatic cancer

Kiemen¹,

Damanakis²,

Braxton³

et al. 2023

Med

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A highly sensitive and specific real-time quantitative PCR for BRAF V600E/K mutation screening

Cited by 8 publications

References 44 publications

Real-Time Polymerase Chain Reaction: Current Techniques, Applications, and Role in COVID-19 Diagnosis

Real-Time Polymerase Chain Reaction: Current Techniques, Applications, and Role in COVID-19 Diagnosis

Solid-Phase Primer Elongation Using Biotinylated dNTPs for the Detection of a Single Nucleotide Polymorphism from a Fingerprick Blood Sample

Tissue clearing and 3D reconstruction of digitized, serially sectioned slides provide novel insights into pancreatic cancer

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