Fluorescent Biosensing Method For Detection Of HIV-related DNA Based On Strand Displacement Amplification And Dnazyme

Human immunodeficiency virus（HIV）can lead to acquired immunodeficiency syndrome（AIDS）by destroying infected patients’immune system.Conventional methods such as enzyme-linked immunosorbent assay（ELISA）and Western blot（WB）are common methods for clinical diagnosis of HIV infection.However,there is a“window period”for HIV detection because it takes a period of time to produce antibody after HIV infected,which makes the early diagnosis of HIV infection full of challenges.Nucleic acid assays have attracted increasing attention because they can prompt for viral infection even in the absence of HIV antibodies.Thus,sensitive and specific detection of HIV-related DNA is of great importance for early accurate diagnosis and therapy of HIV-infected patients.Strand displacement amplification（SDA）is an isothermal nucleic acid amplification strategy with high efficiency.DNAzyme can be applied as signal amplification unit because of its cyclic cleavage property.Therefore,we developed a simple and rapid fluorescence method for HIV-related DNA detection based on SDA and Mg²⁺-dependent DNAzyme reaction.In the presence of target HIV DNA,it can hybridize with template DNA and activate SDA to generate numerous DNAzyme sequences.With introduction of Mg²⁺,DNAzyme can be activated to circularly cleave the substrate DNA,which leads to the separation of fluorophore reporters from the quenchers,resulting in the recovery of the fluorescence.Under the optimal experimental conditions,the established biosensing method can detect target DNA down to 61 fM with a linear range from 100 fM to 1 nM,and discriminate target DNA from mismatched DNA perfectly.In addition,the developed biosensing method was successfully applied to assay target DNA spiked into human serum samples.With the advantages of fast,easy operation and high-performance,this biosensing method might be an alternative tool for clinical diagnosis of HIV infection.