Abstract
A sensitive fluorescent sensor for sequence-specific recognition of double-stranded DNA (dsDNA) was developed on the surface of silver-coated glass slide (SCGS). Oligonucleotide-1 (Oligo-1) was designed to assemble on the surface of SCGS and act as capture DNA, and oligonucleotide-2 (Oligo-2) was designed as signal DNA. Upon addition of target HIV-1 dsDNA (Oligo-3•Oligo-4), signal DNA could bind on the surface of silver-coated glass because of the formation of C•GoC in parallel triplex DNA structure. Biotin-labeled glucose oxidase (biotin-GOx) could bind to signal DNA through the specific interaction of biotin-streptavidin, thereby GOx was attached to the surface of SCGS, which was dependent on the concentration of target HIV-1 dsDNA. GOx could catalyze the oxidation of glucose and yield H2O2, and the HPPA can be oxidized into a fluorescent product in the presence of HRP. Therefore, the concentration of target HIV-1 dsDNA could be estimated with fluorescence intensity. Under the optimum conditions, the fluorescence intensity was proportional to the concentration of target HIV-1 dsDNA over the range of 10 pM to 1000 pM, the detection limit was 3 pM. Moreover, the sensor had good sequence selectivity and practicability and might be applied for the diagnosis of HIV disease in the future.
Highlights
Double-stranded structure, reported in 1953 [1], is the natural state of DNA
Upon addition of target human immunodeficiency virus 1 (HIV-1) double-stranded DNA (dsDNA), glucose oxidase (GOx) could be immobilized on the surface of silver-coated glass slide (SCGS). ereby, the concentration of target HIV-1 dsDNA controlled the number of bound GOx, which could be detected with the fluorescence of oxidized HPPA. ese methods have been applied to develop sensors for sequencespecific recognition of dsDNA based upon triplex formation [13,14,15,16,17]
In order to avoid the nonspeci c adsorption of streptavidin and GOx, DNase I is used to cleave the DNA strand from the surface of SCGS, and the bound glucose oxidase is transferred into the bu er. us, the concentration of target dsDNA is transduced into the concentration of H2O2 which is the oxidative product of glucose in the presence of GOx. en, HPPA can be oxidized into the Capture DNA
Summary
Double-stranded structure, reported in 1953 [1], is the natural state of DNA. double-stranded DNA (dsDNA) detection is of particular importance in gene therapy, diagnosis, and monitoring fatal infections caused by viruses and diseases that are associated with genetic alterations [2,3,4,5,6,7,8]. Routine protocols for sequence-specific recognition of dsDNA, for instance, are performed by using zinc finger DNA-binding proteins [9, 10], polyamides [11, 12], and triplex-forming oligonucleotides [13,14,15,16,17]. Enzymes are highly applicable in biosensors as recognition and signaling elements for the detection of specific molecules due to the features such as high sensitivity and good selectivity [19]. Given these favorable characteristics, glucose oxidase (GOx) can catalyze the oxidation of glucose and is one of the cheapest and most stable redox enzymes. GOx is conjugated for recognition of biomolecules and acted as an amplifying label, which is successfully applied to establish sensors for proteins [29, 30] and DNA [31]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
