CRISPR-Based Approaches for Efficient and Accurate Detection of SARS-CoV-2

Wancun Zhang, PhD; Kangbo Liu, MD; Pin Zhang, MD; Weyland Cheng, PhD; Linfei Li, MD; Fan Zhang, MD; Zhidan Yu, PhD; Lifeng Li, PhD; Xianwei Zhang, PhD

Disclosures

Lab Med. 2021;52(2):116-121. 

In This Article

Efficient and Accurate Detection of SARS-CoV-2

Rapid, effcient, and accurate identification of infectious diseases is essential to optimize clinical care and guide infection control and public health interventions to limit disease spread in both highly specialized medical centers and remote health care settings. Many methods exist for detecting nucleic acids, and each technology has different advantages and limitations.[14,18–22] The ideal diagnostic test would be inexpensive and accurate and would provide a result rapidly, allowing for point-of-care use on multiple specimen types without the need for technical personnel or sophisticated equipment. Highly pathogenic viruses can emerge in remote settings but can also spread globally (eg, Ebola virus and Middle East respiratory syndrome coronavirus), requiring a method that provides early rapid and accurate detection, limiting the spread of infectious diseases and promoting timely care.[23]

The CRISPR and CRISPR-associated (Cas) adaptive immune systems contain programmable endonucleases that can be used for CRISPR-based diagnostics. Although some Cas enzymes target DNA, single-effector RNA-guided RNases, such as Cas13a, can be reprogrammed with CRISPR RNAs (crRNAs) to provide a platform for specific RNA sensing. Upon recognition of its RNA target, activated Cas13a engages in "collateral" cleavage of nearby nontargeted RNAs, which allows Cas13a to detect the presence of a specific RNA in vivo by triggering programmed cell death or in vitro by nonspecific degradation of labeled RNA. The Specific High Sensitivity Enzymatic Reporter Unlocking (SHERLOCK), based on nucleic acid amplification and Cas13a-mediated collateral cleavage of a reporter RNA, allows for real-time, rapid, and specific detection of the target with attomolar (aM) sensitivity.[17,24]

Compared with the RT-PCR-based approach, CRISPR-based approaches have the following advantages: isothermal signal amplification obviating the need for thermocycling, rapid turnaround time, single nucleotide target specificity, integration with accessible and easy-to-use reporting formats such as lateral flow strips, and no requirements for complex laboratory infrastructure.[16] Therefore, CRISPR-based approaches are expected to be used for the rapid, sensitive, and visual detection of SARS-CoV-2.

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