Engineering a DNA-cleaving DNAzyme and PCR into a simple sensor for zinc ion detection

Abstract

The development of a simple sensor (9NL27-Zn) based on DNAzyme and PCR and aimed at the detection of low concentrations of zinc (II) ions is described. A specific Zn(II)-dependent DNAzyme (9NL27) with DNA-cleaving activity was employed. In the presence of zinc (II), the DNAzyme hydrolyzed DNA substrate into two pieces (5' and 3' fragments), forming 3'-terminal hydroxyl in the 5' fragment and 5'-phosphate in the 3' fragments. Subsequently, the 5' fragment left the DNAzyme and bound a short DNA template. The 5' fragment was used as a primer and extended a single-stranded full-length template by Taq polymerase. Finally, this full-length template was amplified by PCR. The amplified products had a quantitative relationship with Zn(II) concentration. Under our experimental conditions, the DNA sensor showed sensitivity (10 nM) and high specificity for zinc ion detection. After improvement of the DNA sensor, the detection limit can reach 1 nM. The simple DNA sensor may become a DNA model for the detection of trace amounts of other targets.

Zn2+‐Dependent DNAzymes: From Solution Chemistry to Analytical, Materials and Therapeutic Applications

“…[38] This DNAzyme was later used as a sensor for Zn 2 + detection. [94] Gu and Breaker reported a new selection method using a circular library and obtained a few small DNA-cleaving DNAzymes (Figure 5A). [37] IÀ R1 is a self-cleaving DNAzyme, while IÀ R3 can be separated into a substrate strand and an enzyme strand.…”

“…Later, the group found that by just mutating two nucleotides in the original 10MD5, the requirement of Mn 2+ can be omitted and activity was achieved with Zn 2+ alone [38] . This DNAzyme was later used as a sensor for Zn 2+ detection [94] …”

“…Taking advantage of the adsorption property of TiS 2 nanosheets, a fluorescently labeled DNAzyme was used for detecting Zn 2+ by measuring fluorescence polarization, [106,107] although the quenching effect of the material was not considered for the quantification [108] . Zn 2+ detection was also examined using the 9NL27 DNAzyme, which is a variant of 10MD5 [94] . This DNAzyme contains a longer catalytic core compared to I−R3, but a similar limit of detection of 1 nM Zn 2+ was also reached.…”

Simulation/Experiment Confrontation, an Efficient Approach for Sensitive SAW Sensors Design

“…The choice of these ions is related to their presence in Sarthe River Basin of "Le Mans" region [32]. Several receptors have been investigated for zinc ions detection, mainly ion imprinted polymers [33][34][35], DNA [36], aptamers [37], self-assembled monolayers (SAM) [38,39], enzymes [40,41], and molecular cages [42][43][44][45]. Herein, we have chosen to functionalize the sensitive zone of the 104 MHz and 208 MHz SAW sensor with 9-{[4-({[4-(9 anthrylmethoxy) phenyl]sulfanyl} methyl)]methyl] anthracene (TDP-AN).…”

Single‐gap Microelectrode Functionalized with Single‐walled Carbon Nanotubes and Pbzyme for the Determination of Pb2+

“…It can bind with metal ion specifically in a short time, and then cleave the complementary single‐stranded DNA (CS‐DNA) at a specific site , which can overcome the long analysis time using aptamer‐based biosensors, and poor stability of protease‐based biosensors. Up to now, metal ions including Na + , Ag + , Hg 2+ , Pb 2+ , Cu 2+ , Ca 2+ , Zn 2+ , Cd 2+ , Cr 3+ have been detected using DNAzymes . A variety of DNAzyme‐modified biosensors for Pb 2+ detection were reported.…”