Detection of Single Nucleotide Polymorphisms by Fluorescence Embedded Dye SYBR Green I Based on Graphene Oxide

Abstract

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The detection of single nucleotide polymorphisms (SNPs) is of great significance in the early diagnosis of diseases and the rational use of drugs. Thus, a novel biosensor based on the quenching effect of fluorescence-embedded SYBR Green I (SG) dye and graphene oxide (GO) was introduced in this study. The probe DNA forms a double helix structure with perfectly complementary DNA (pcDNA) and 15 single-base mismatch DNA (smDNA) respectively. SG is highly intercalated with perfectly complementary dsDNA (pc-dsDNA) and exhibits strong fluorescence emission. Single-base mismatch dsDNA (SNPs) has a loose double-stranded structure and exhibits poor SG intercalation and low fluorescence sensing. At this time, the sensor still showed poor SNP discrimination. GO has a strong effect on single-stranded DNA (ssDNA), which can reduce the fluorescence response of probe DNA and eliminate background interference. And competitively combined with ssDNA in SNPs, quenching the fluorescence of SG/SNP, while the fluorescence value of pc-dsDNA was retained, increasing the signal-to-noise ratio. At this time, the sensor has obtained excellent SNP resolution. Different SNPs detect different intensities of fluorescence in the near-infrared region to evaluate the sensor's identification of SNPs. The experimental parameters such as incubation time, incubation temperature and salt concentration were optimized. Under optimal conditions, 1 nM DNA with 0–10 nM linear range and differentiate 5% SNP were achieved. The detection method does not require labeling, is low cost, simple in operation, exhibits high SNP discrimination and can be distinguished by SNP at room temperature.

Keywords

• graphene oxide
• single nucleotide polymorphism
• SYBR Green I
• DNA sensor
• fluorescence