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

Jiaoyun Xia,Jing Qing,Lihua Wang,Junlong Tang,Tong Xu

doi:10.3389/fchem.2021.631959

Abstract

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.

Highlights

Single nucleotide polymorphisms (SNPs) are extremely important genetic variations in human genes (Martin et al, 2020)
A novel biosensor based on the quenching effect of fluorescence-embedded SYBR Green I (SG) dye (Hur et al, 2019) and graphene oxide (GO) (Li et al, 2013; Wan et al, 2017; Jiang et al, 2020) was constructed
SG is well embedded in fully complementary dsDNA and produces high fluorescence

Summary

Introduction

Single nucleotide polymorphisms (SNPs) are extremely important genetic variations in human genes (Martin et al, 2020). DNA Sensor technology (Tu et al, 2018), etc These methods can effectively detect SNPs, most of them PCR technology is required to amplify DNA (Qing et al, 2019), which is expensive and timeconsuming. A fast, economical and reliable SG embedded sensor is introduced to detect SNP. Such as, inserting SG into DNA double strands, and immobilizing DNA on the surface of molybdenum disulfide-gold nanoparticles (MoS2@AuNPs), and MoS2@AuNPs quenches the fluorescence of SG inserted into the DNA double strands. Only with the participation of SG, it is difficult for the sensor to achieve high resolution and high accuracy SNP detection

Methods

Results

Conclusion