Novel nucleic acid detection strategies based on CRISPR‐Cas systems: From construction to application

Zhu, Chushu; Liu, Chuanyang; Xie, Shuhong; Li, Wen‐ying; Zhu, Lingyun; Zhu, Lv‐yun

doi:10.1002/bit.27334

Cited by 45 publications

(29 citation statements)

References 95 publications

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“…The dCas9 system is designed with modification in the basic activity of the conventional Cas9 protein in terms of deactivating the nucleic acid cleavage potential and only retaining the specific binding ability to target dsDNA. This was accomplished by inducing two point mutations H840A and D10A in the HNH and RuvC nuclease domain of the conventional Cas9 effector protein [36,53].…”

Section: Crispr In Diagnosismentioning

confidence: 99%

“…In contrast to this, Cas13a, which is previously referred to as C2c2 are RNA-guided targeting enzymes involved in type VI of CRISPR/Cas system and are specific for ssRNA. Based on the promiscuous Rnase ability of collateral cleavage of Cas12a as well as Cas13a several molecular diagnostic platforms have been developed in the recent years details of which will be discussed further [53][54][55][56][57].…”

Section: Crispr In Diagnosismentioning

confidence: 99%

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CRISPR Cas/Exosome Based Diagnostics: Future of Early Cancer Detection

Mubthasima¹,

Pande²,

Prakash³

et al. 2022

Rural Health

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Trending and Thriving, CRISPR/Cas has expanded its wings towards diagnostics in recent years. The potential of evading off targeting has not only made CRISPR/Cas an effective therapeutic aid but also an impressive diagnostic tool for various pathological conditions. Exosomes, 30 - 150nm sized extracellular vesicle present and secreted by almost all type of cells in body per se used as an effective diagnostic tool in early cancer detection. Cancer being the leading cause of global morbidity and mortality can be effectively targeted if detected in the early stage, but most of the currently used diagnostic tool fails to do so as they can only detect the cancer in the later stage. This can be overcome by the use of combo of the two fore mentioned diagnostic aids, CRISPR/Cas alongside exosomes, which can bridge the gap compensating the cons. This chapter focus on two plausible use of CRISPR/Cas, one being the combinatorial aid of CRISPR/Cas and Exosome, the two substantial diagnostic tools for successfully combating cancer and other, the use of CRISPR in detecting and targeting cancer exosomes, since they are released in a significant quantity in early stage by the cancer cells.

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Section: Crispr In Diagnosismentioning

confidence: 99%

Section: Crispr In Diagnosismentioning

confidence: 99%

CRISPR Cas/Exosome Based Diagnostics: Future of Early Cancer Detection

Mubthasima¹,

Pande²,

Prakash³

et al. 2022

Rural Health

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“…In recent years, clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR associated (Cas) enzymes have been playing increasingly important roles part in both genome editing and nucleic acid detection (Zhu et al, 2020). For instance, with the mediation of CRISPR RNA (crRNA), Cas12a can recognize specific DNA sequences and exhibit nonspecific single‐stranded DNA (ssDNA) cleavage activity (Swarts & Jinek, 2019; Yamano et al, 2016).…”

Section: Introductionmentioning

confidence: 99%

A CRISPR‐Cas12a‐derived biosensor enabling portable personal glucose meter readout for quantitative detection of SARS‐CoV‐2

Huang

Shi

Qian

et al. 2021

Biotech & Bioengineering

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Severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) has spread rapidly throughout the whole world and caused significant difficulties in the prevention and control of the epidemic. In this case, several detection methods have been established based on nucleic acid diagnostic techniques and immunoassays to achieve sensitive and specific detection of SARS‐CoV‐2. However, most methods are still largely dependent on professional instruments, highly trained operators, and centralized laboratories. These limitations gravely diminish their practicality and portability. Herein, a clustered regularly interspaced short palindromic repeats (CRISPR) Cas12a based assay was developed for portable, rapid and sensitive of SARS‐CoV‐2. In this assay, samples were quickly pretreated and amplified by reverse transcription recombinase‐aided amplification under mild conditions. Then, by combining the CRISPR Cas12a system and a glucose‐producing reaction, the signal of the virus was converted to that of glucose, which can be quantitatively read by a personal glucose meter in a few seconds. Nucleocapsid protein gene was tested as a model target, and the sensitivity for quantitative detection was as low as 10 copies/μl, which basically meet the needs of clinical diagnosis. In addition, with the advantages of lower material cost, shorter detection time, and no requirement for professional instrument in comparison with quantitative reverse transcription‐polymerase chain reaction, this assay is expected to provide a powerful technical support for the early diagnosis and intervention during epidemic prevention and control.

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“…CRISPR-Cas systems are widely used for genome editing [ 20 ] and, in recent years, the trans-cleavage activity of Cas proteins has been discovered [ 21 ]. Detection methods based on CRISPR-Cas, which is now also considered a next-generation pathogen detection method, have become established [ 22 , 23 ]. DETECTR (DNA endonuclease targeted CRISPR trans-reporter) [ 24 ] and HOLMES (one-HOur Low-cost Multipurpose highly Efficient System) [ 25 ] can detect DNA sequences with attomolar sensitivity and high specificity using CRISPR-Cas12a [ 26 ].…”

Section: Introductionmentioning

confidence: 99%

Obtaining Specific Sequence Tags for Yersinia pestis and Visually Detecting Them Using the CRISPR-Cas12a System

Chen¹,

Lyu²,

Wang³

et al. 2021

Pathogens

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Three worldwide historical plague pandemics resulted in millions of deaths. Yersinia pestis, the etiologic agent of plague, is also a potential bioterrorist weapon. Simple, rapid, and specific detection of Y. pestis is important to prevent and control plague. However, the high similarity between Y. pestis and its sister species within the same genus makes detection work problematic. Here, the genome sequence from the Y. pestis CO92 strain was electronically separated into millions of fragments. These fragments were analyzed and compared with the genome sequences of 539 Y. pestis strains and 572 strains of 20 species within the Yersinia genus. Altogether, 97 Y. pestis-specific tags containing two or more single nucleotide polymorphism sites were screened out. These 97 tags efficiently distinguished Y. pestis from all other closely related species. We chose four of these tags to design a Cas12a-based detection system. PCR–fluorescence methodology was used to test the specificity of these tags, and the results showed that the fluorescence intensity produced by Y. pestis was significantly higher than that of non-Y. pestis (p < 0.0001). We then employed recombinase polymerase amplification and lateral flow dipsticks to visualize the results. Our newly developed plasmid-independent, species-specific library of tags completely and effectively screened chromosomal sequences. The detection limit of our four-tag Cas12a system reached picogram levels.

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Novel nucleic acid detection strategies based on CRISPR‐Cas systems: From construction to application

Cited by 45 publications

References 95 publications

CRISPR Cas/Exosome Based Diagnostics: Future of Early Cancer Detection

CRISPR Cas/Exosome Based Diagnostics: Future of Early Cancer Detection

A CRISPR‐Cas12a‐derived biosensor enabling portable personal glucose meter readout for quantitative detection of SARS‐CoV‐2

Obtaining Specific Sequence Tags for Yersinia pestis and Visually Detecting Them Using the CRISPR-Cas12a System

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