Csm6-DNAzyme Tandem Assay for One-Pot and Sensitive Analysis of Lead Pollution and Bioaccumulation in Mice

Abstract

Lead contamination in the environment tends to enter the food chain and further into the human body, causing serious health issues. Herein, we proposed a Csm6-DNAzyme tandem assay (termed cDNAzyme) using CRISPR/Cas III-A Csm6 and GR-5 DNAzyme, enabling one-pot and sensitive detection of lead contamination. We found that Pb2+-activated GR-5 DNAzyme produced cleaved substrates that can serve as the activator of Csm6, and the Csm6-DNAzyme tandem improved the sensitivity for detecting Pb2+ by 6.1 times compared to the original GR-5 DNAzyme. Due to the high specificity of DNAzyme, the cDNAzyme assay can discriminate Pb2+ from other bivalent and trivalent interfering ions and allowed precise detection of Pb2+ in water and food samples. Particularly, the assay can achieve one-step, mix-and-read detection of Pb2+ at room temperature. We used the cDNAzyme assay to investigate the accumulation of lead in mice, and found that lead accumulated at higher levels in the colon and kidney compared to the liver, and most of the lead was excreted. The cDNAzyme assay is promising to serve as analytical tools for lead-associated environmental and biosafety issues.

Peroxidase-Mimicking DNAzymes as Receptors for Label-Free Discriminating Heavy Metal Ions by Chemiluminescence Sensor Arrays

“…Heavy metal ions are mostly produced during industrial production. Because it is difficult to be biodegraded, it accumulates in the body through the transmission of the food chain, causing chronic poisoning, thus endangering human health and destroying the ecosystem. − For example, mercury ions inhibit enzyme activity and cause cellular damage, disrupting the central nervous and endocrinium. Ingestion of chromium ions into the human body can cause skin ulcers, impaired liver function, and lung cancer.…”

CRISPR-based nucleic acid diagnostics for pathogens

“…Besides, harnessing the changes in pH and pyrophosphate concentration that occur during cyclic nucleic polymerization could realize the colorimetric or visible fluorometric detection of SARS-CoV-2 [ 38 ]. Inspired by the use of activator variant that cannot be degraded for TtCsm6 [ 67 , 92 ], Wiedenheft group adopted the Can2 nuclease to assist the signaling of subtype III-A CRISPR capture technique [ 41 ]. Use of Can2 lacking the ring nuclease activity can sustain a high level of cA 4 activator, contributing to an sensitivity improvement.…”

CRISPR/Cas System: The Accelerator for the Development of Non-nucleic Acid Target Detection in Food Safety

“…The detection sensitivity of Pb 2+ was enhanced 6.1-fold compared with the traditional GR-5 DNAzyme. 47 3.2. Detection of ATP.…”

“…Then the generated products could act as the activator of Csm6 for subsequent reactions. The detection sensitivity of Pb 2+ was enhanced 6.1-fold compared with the traditional GR-5 DNAzyme …”

“…(C) Exonuclease III-assisted Cas12a electrochemiluminescence biosensor for mercury determination 46. (D) Csm6-DNAzyme tandem assay for lead analysis 47.…”