期刊
ACS SENSORS
卷 7, 期 2, 页码 453-459出版社
AMER CHEMICAL SOC
DOI: 10.1021/acssensors.1c02113
关键词
conformational polymorphism; diagnosis; G-quadruplex; fluorescence; SARS-CoV-2
资金
- JNCASR [JNC/JNCASR/D0018/2021/01178]
- BRICS Multilateral RD Project [DST/IMRCD/BRIC/PilotCAll2/EPNAPT/2018(G)]
- Indo-French project [IFCPAR/CEFIPRA-62T10-3]
- Department of Science and Technology (DST), Govt. of India
- Wellcome Trust/DBT India Alliance Grant [IA/S/16/2/502700]
- DBT-IISc Partnership Program [22-0905-0006-05-987-436]
- UGC
- IISc
Identifying and characterizing a highly conserved antiparallel G-quadruplex (GQ)-forming DNA sequence within the SARS-CoV-2 genome, researchers developed a novel benzobisthiazole-based fluorogenic probe for specific target recognition. This probe, used in a cost-effective assay involving PCR amplification and endpoint fluorescence detection, demonstrates a potential diagnostic tool for SARS-CoV-2 clinical samples. The study suggests a general strategy that can be adapted for developing specific diagnostic assays targeting different noncanonical nucleic acid sequences.
Unravelling unique molecular targets specific to viruses is challenging yet critical for diagnosing emerging viral diseases. Nucleic acids and proteins are the major targets in diagnostic assays of viral pathogens. Identification of novel sequences and conformations of nucleic acids as targets is desirable for developing diagnostic assays specific to a virus of interest. Here, we disclose the identification and characterization of a highly conserved antiparallel G-quadruplex (GQ)-forming DNA sequence present within the SARS-CoV-2 genome. The two-quartet GQ with unique loop compositions formed a distinct recognition motif. Design, synthesis, and fine tuning of structure-activity of a set of small molecules led to the identification of a benzobisthiazole-based fluorogenic probe which unambiguously recognizes the target SARS-CoV-2 GQ DNA. A robust cost-effective assay was developed through thermal cycler PCR-based amplification of the antiparallel GQ-forming ORF1ab region of the SARS-CoV-2 genome and endpoint fluorescence detection with the probe. An exclusive pH window (3.5-4) helped trigger reliable conformational polymorphism (RCP) involving DNA duplex to GQ transformation, which aided the development of a GQ-RCP platform for the diagnosis of SARS-CoV-2 clinical samples. This general strategy can be adapted for the development of specific diagnostic assays targeting different noncanonical nucleic acid sequences.
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