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Nucleotide-selective amplification and array-based detection for identifying multiple somatic mutations
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DOI：10.1016/j.aca.2023.341343.png)
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Abstract
En 中文
In the context of personalized and cost-effective treatment, knowledge of the mutational status of specific genes is advantageous to predict which patients are responsive to therapies. As an alternative to one-by-one detection or massive sequencing, the presented genotyping tool determines multiple polymorphic sequences that vary a single nucleotide. The biosensing method includes an effective enrichment of mutant variants and selective recognition by colorimetric DNA arrays. The proposed approach is the hybridization between sequence-tailored probes and products from PCR with SuperSelective primers to discriminate specific variants in a single locus. A fluorescence scanner, a documental scanner, or a smartphone captured the chip images to obtain spot intensities. Hence, specific recognition patterns identified any single-nucleotide change in the wild-type sequence over- coming qPCR methods and other array-based approaches. Studied mutational analyses applied to human cell lines provided high discrimination factors, the precision was 95%, and the sensitivity was 1% mutant of total DNA. Also, the methods showed a selective genotyping of the KRAS gene from tumorous samples (tissue and liquid biopsy), corroborating results by NGS. The developed technology supported on low-cost robust chips and optical reading provides an attractive pathway toward implementing fast, cheap, reproducible discrimination of oncological patients.
Keywords:
Array sensing
DNA chip technology
Allele-specific techniques
Mutations in oncogenes
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