Tb^(3+)-nucleic acid probe-based label-free and rapid detection of mercury pollution in food

Mercury is a threatening pollutant in food,herein,we developed a Tb^(3+)-nucleic acid probe-based label-free assay for mix-and-read,rapid detection of mercury pollution.The assay utilized the feature of light-up fluorescence of terbium ions(Tb^(3+))via binding with single-strand DNA.Mercury ion,Hg^(2+)induced thymine(T)-rich DNA strand to form a double-strand structure(T-Hg^(2+)-T),thus leading to fluorescence reduction.Based on the principle,Hg^(2+)can be quantified based on the fluorescence of Tb^(3+),the limit of detection was 0.0689μmol/L and the linear range was 0.1-6.0μmol/L.Due to the specificity of T-Hg^(2+)-T artificial base pair,the assay could distinguish Hg^(2+)from other metal ions.The recovery rate was ranged in 98.71%-101.34%for detecting mercury pollution in three food samples.The assay is low-cost,separation-free and mix-to-read,thus was a competitive tool for detection of mercury pollution to ensure food safety.

Real-time in situ observation of P53-mediated cascade activation of apoptotic pathways with nucleic acid multicolor fluorescent probes based on symmetrical gold nanostars

T-2 toxin,one of the most dangerous natural pollutants,induces apoptosis through multiple pathways.Amongst,P53 mediated apoptosis pathway,an important collection of molecules,plays a key role in cell vital activity.Real-time monitoring of upstream and downstream activation relationships of P53 mRNA,Bax mRNA,and cytochrome c(Cyt c)in signaling pathways is of great significance for understanding the apoptotic machinery in human physiology.In this work,a novel nucleic acid multicolor fluorescent probe,based on silica-coated symmetric gold nanostars(S-AuNSs@SiO_(2)),was developed for highly sensitive in situ real-time imaging of P53 mRNA,Bax mRNA,and Cyt c during T-2 toxin-induced apoptosis.The nucleic acid chains modified with carboxyl groups were modified on the surface of S-AuNSs@SiO_(2)by amide reaction.The complementary chains of targeted mRNA and the aptamer of targeted Cyt c were modified with different fluorophores,respectively,and successfully hybridized on S-AuNSs@SiO_(2)surface.When targets were present,the fluorescent chains bound to the targets and detached from the material,resulting in the quenched fluorescence being revived.The probes based on S-AuNSs showed excellent performance is partly ascribed to the presence of 20 symmetric“hot spots”.Notably,the amide-bonded probe exhibited excellent anti-interference capability against biological agents(nucleases and biothiols).During the real-time fluorescence imaging of T-2 toxin-induced apoptosis,the corresponding fluorescence signals of P53 mRNA,Bax mRNA,and Cyt c were observed sequentially.Therefore,S-AuNSs@SiO_(2)probe not only provides a novel tool for real-time monitoring of apoptosis pathways cascade but also has considerable potential in disease diagnosis and pharmaceutical medical.

Thermodynamics-guided two-way interlocking DNA cascade system for universal multiplexed mutation detection

Detection of point mutations in driver genes is of great significance for the early diagnosis,treatment,and prognostic evaluation of cancer.However,current detection methods do not offer versatility,specificity,and rapid performance simultaneously.Thus,multiple mutation detection processes are necessary,which results in long processing times and high costs.In this study,we developed a thermodynamics-guided two-way interlocking DNA cascade system for universal multiplexed mutation detection(TTI-CS).This strategy is based on the DNA probe,which changes the thermodynamic balance of the DNA cascade by the designed bubble structure,thereby achieving a good distinction between mutant and wild-type DNA.The designed method greatly shortens the detection time through two-way intrusion.In addition,this method only changes two inexpensive trigger and bridge sequences,which replace the specific and expensive nucleic acid probes used in analyses based on traditional DNA probe methods,thereby enabling multiple detections.We performed the detection of synthetic single-stranded DNA for the five mutation points and successfully detected in endometrial cancer specimens.The detection limit of this method is0.1%,which better meets the needs of clinical low-abundance multiple mutation detection.Overall,TTI-CS is currently one of the best methods for detecting multiple mutation detections.

Shared-probe system:An accurate,low-cost and general enzyme-assisted DNA probe system for detection of genetic mutation

Enzyme assisted DNA probes are powerful tools in molecular diagnostics for their simplicity,rapidity,and low detection limit.However,cost of probes,difficulty in optimization and disturbance of secondary structure hindered the wider application of enzyme assisted DNA probes.To solve the problems,we designed a new system named shared-probe system.By introducing two unlabeled single stranded DNA named Sh1 and Sh2 as the bridge between probe and the substrate,the same sequence of dually labeled probe with stable performance was shared for different mutations,thus sparing the expense and time cost on designing,synthesizing and optimizing corresponding probes.Besides,the hybridization between Sh1 and the substrate could overcome secondary structures,which guaranteed the detection of different substrates.The performance and generality of the design were tested by low abundance detection in synthetic single DNA samples and the limit of detection was 0.05%for PTENR130 Q,EGFR-L858 R and 0.02%for BRCA1-NM007294.3.In genomic DNA samples,the limit of detection of 0.1%can be achieved for EGFR-L858 R,demonstrating the potential of clinical application in our design.