Digital CRISPR-based method for the rapid detection and absolute quantification of nucleic acids

Wu, Xiaolin; Tay, Joshua K.; Goh, Chuan Keng; Chan, Cheryl; Lee, Yie Hou; Springs, Stacy L.; Wang, De Yun; Loh, Kwok Seng; Lu, Timothy K.; Yu, Hanry

doi:10.1016/j.biomaterials.2021.120876

Cited by 66 publications

(44 citation statements)

References 73 publications

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“…Our preliminary results indicated that CRISPR-Cas12a direct detection could detect ∼10 9 copies of plasmid DNA per reaction in under 30 min of measurement time. Previous studies also indicated a low detection limit of 10 pM target DNAs in a direct Cas12a assay ( 52 , 53 ) or fM-level RNA in Cas13a assay ( 32 ). To improve the detection limit, Yue et al described a droplet Cas amplification-free assay in picoliter-sized droplets to increase local molecule concentration and enhance reaction efficiency ( 54 ).…”

Section: Discussionmentioning

confidence: 94%

CRISPR-Cas12a-Based Detection for the Major SARS-CoV-2 Variants of Concern

et al. 2021

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We described CRISPR-Cas12-based multiplex allele-specific assay for rapid SARS-CoV-2 variant genotyping. The new system has the potential to be quickly developed, continuously updated, and easily implemented for screening of SARS-CoV-2 variants in resource-limited settings. This approach can be adapted for emerging mutations and implemented in laboratories already conducting SARS-CoV-2 nucleic acid amplification tests using existing resources and extracted nucleic acid.

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Section: Discussionmentioning

confidence: 94%

CRISPR-Cas12a-Based Detection for the Major SARS-CoV-2 Variants of Concern

et al. 2021

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“…The combination of isothermal amplification and CRISPR-Cas12-based nucleic acids detection achieves a second recognition step for amplification products, which has the potential to build a rapid, sensitive and accurate quantitative detection platform. Digitization-enhanced CRISPR/Cas-assisted one-pot virus detection (deCOViD) and rapid digital CRISPR approach (RADICA) were two quantitative detection technologies taking advantage of AIOD-CRISPR [ 55 , 56 ] (Fig. 3 a).…”

Section: Improvement Of Quantitative Detection Capabilitiesmentioning

confidence: 99%

“…3 a). However, in order to avoid undesired premature target amplification at room temperature, quantitative detection based on RPA needs to prepare all reaction mixture on ice and loads into the chip within 1 min after adding Mg 2+ , which makes the detection procedure more complex [ 56 ]. Therefore, Ding et al [ 38 ] solved this problem by increasing the initiation temperature of amplification, and constructed digital warm-start CRISPR (dWS-CRISPR) based on one-pot WS-CRISPR (Fig.…”

Section: Improvement Of Quantitative Detection Capabilitiesmentioning

confidence: 99%

Improved Strategies for CRISPR-Cas12-based Nucleic Acids Detection

2022

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The COVID-19 pandemic has brought great challenges to traditional nucleic acid detection technology. Thus, it is urgent to develop a more simple and efficient nucleic acid detection technology. CRISPR-Cas12 has signal amplification ability, high sensitivity and high nucleic acid recognition specificity, so it is considered as a nucleic acid detection tool with broad development prospects and high application value. This review paper discusses recent advances in CRISPR-Cas12-based nucleic acid detection, with an emphasis on the new research methods and means to improve the nucleic acid detection capability of CRISPR-Cas12. Strategies for improving sensitivity, optimization of integrated detection, development of simplified detection mode and improvement of quantitative detection capabilities are included. Finally, the future development of CRISPR-Cas12-based nucleic acids detection is prospected.

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“…By using the CRISPR/Cas9 system, clearance of viruses from infected cells becomes hypothetically practical for any DNA- or RNA-mediated virus during their pathological process. Therefore, the CRISPR/Cas9 technique has become a game-changing tool for modifying several developmental phases of the viral life cycle and holds the capacity to enable efficient genetic therapy versus human viruses (Table 2 ) [ 43 , 44 ]. In recent years, CRISPR/Cas9-mediated antiviral protocols to manipulate infectious human viruses have been applied efficiently.…”

Section: Crispr/cas9 Applications In Viral Infections and Genetic Dis...mentioning

confidence: 99%

CRISPR/Cas9 application in cancer therapy: a pioneering genome editing tool

et al. 2022

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The progress of genetic engineering in the 1970s brought about a paradigm shift in genome editing technology. The clustered regularly interspaced short palindromic repeats/CRISPR associated protein 9 (CRISPR/Cas9) system is a flexible means to target and modify particular DNA sequences in the genome. Several applications of CRISPR/Cas9 are presently being studied in cancer biology and oncology to provide vigorous site-specific gene editing to enhance its biological and clinical uses. CRISPR's flexibility and ease of use have enabled the prompt achievement of almost any preferred alteration with greater efficiency and lower cost than preceding modalities. Also, CRISPR/Cas9 technology has recently been applied to improve the safety and efficacy of chimeric antigen receptor (CAR)-T cell therapies and defeat tumor cell resistance to conventional treatments such as chemotherapy and radiotherapy. The current review summarizes the application of CRISPR/Cas9 in cancer therapy. We also discuss the present obstacles and contemplate future possibilities in this context.

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Digital CRISPR-based method for the rapid detection and absolute quantification of nucleic acids

Cited by 66 publications

References 73 publications

CRISPR-Cas12a-Based Detection for the Major SARS-CoV-2 Variants of Concern

CRISPR-Cas12a-Based Detection for the Major SARS-CoV-2 Variants of Concern

Improved Strategies for CRISPR-Cas12-based Nucleic Acids Detection

CRISPR/Cas9 application in cancer therapy: a pioneering genome editing tool

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