ArticlePloS one2015
Single-Tubed Wild-Type Blocking Quantitative PCR Detection Assay for the Sensitive Detection of Codon 12 and 13 KRAS Mutations.
Article in PloS one, 2015. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 8 papers.
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Who cites it
8 citing papers in PubMed, 11 citations in OpenAlex.
- A Novel Self-Competitive Fishing Primer qPCR Approach for Efficient POLE Mutation Detection in Endometrial Cancer Molecular Classification.Current issues in molecular biology · 2026Article
- Combining E-ice-COLD-PCR and Pyrosequencing with Di-Base Addition (PDBA) Enables Sensitive Detection of Low-Abundance Mutations.Applied biochemistry and biotechnology · 2024Article
- Allele-Specific PCR forDiagnostics (Basel, Switzerland) · 2020Article
- PNA Clamping in Nucleic Acid Amplification Protocols to Detect Single Nucleotide Mutations Related to Cancer.Molecules (Basel, Switzerland) · 2020Review
- Sensitive detection of low-abundance in-frame deletions in EGFR exon 19 using novel wild-type blockers in real-time PCR.Scientific reports · 2019Article
- Improved detection of BRAF V600E using allele-specific PCR coupled with external and internal controllers.Scientific reports · 2017Article
- Rapid and accurate detection ofOncotarget · 2017Article
- Wild‑type blocking pcr coupled with internal competitive amplified fragment improved the detection of rare mutation of KRAS.Molecular medicine reports · 2017Article
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Authors and funding
10 authors at 2 institutions in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
The high degree of intra-tumor heterogeneity has meant that it is important to develop sensitive and selective assays to detect low-abundance KRAS mutations in metastatic colorectal carcinoma (mCRC) patients. As a major potential source of tumor DNA in the aforementioned genotyping assays, it was necessary to conduct an analysis on both the quality and quantity of DNA extracted from formalin-fixed paraffin-embedded (FFPE). Therefore, four commercial FFPE DNA extraction kits were initially compared with respect to their ability to facilitate extraction of amplifiable DNA. The results showed that TrimGen kits showed the greatest performance in relation to the quality and quantity of extracted FFPE DNA solutions. Using DNA extracted by TrimGen kits as a template for tumor genotyping, a real-time wild-type blocking PCR (WTB-PCR) assay was subsequently developed to detect the aforementioned KRAS mutations in mCRC patients. The results showed that WTB-PCR facilitated the detection of mutated alleles at a ratio of 1:10,000 (i.e. 0.01%) wild-type alleles. When the assay was subsequently used to test 49 mCRC patients, the results showed that the mutation detection levels of the WTB-PCR assay (61.8%; 30/49) were significantly higher than that of traditional PCR (38.8%; 19/49). Following the use of the real-time WTB-PCR assay, the ΔCq method was used to quantitatively analyze the mutation levels associated with KRAS in each FFPE sample. The results showed that the mutant levels ranged from 53.74 to 0.12% in the patients analyzed. In conclusion, the current real-time WTB-PCR is a rapid, simple, and low-cost method that permits the detection of trace amounts of the mutated KRAS gene.
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