The applications of CRISPR/Cas system in molecular detection

Abstract

The Streptococcus pyogenes CRISPR/Cas system has found widespread applications as a gene‐editing and regulatory tool for the simultaneous delivery of the Cas9 protein and guide RNAs into the cell, thus making the recognition of specific DNA sequences possible. The recent study that shows that Cas9 can also bind to and cleave RNA in an RNA‐programmable manner is suggestive of potential utility of this system as a universal nucleic‐acid recognition tool. To increase the signal intensity of the CRISPR/Cas system, a signal amplification technique has to be exploited appropriately; this requirement is also a challenge for the detection of DNA or RNA. Furthermore, the CRISPR/Cas system may be used to detect point mutations or single‐nucleotide variants because of the specificity of the recognition between the target sequence and the CRISPR/Cas system. These lines of evidence make this technique capable of detecting pathogens during infection via analysis of their DNA or RNA. Thus, here we summarize applications of the CRISPR/Cas system to the recognition and detection of DNA and RNA molecules as well as the signal amplification. We also describe its potential ability to detect mutations and single‐nucleotide variants. Finally, we sum up its applications to testing for pathogens and potential barriers for its implementation.

Discovery and development of safe-in-man broad-spectrum antiviral agents

“…RFPexpressing RVFV, nanoLuc-expressing CHIKV and RRV, as well as GFP-expressing FLUAV, HCV and HMPV also allowed identification of novel activities of several BSAAs (Andersen et al, 2019b;Bosl et al, 2019;de Graaf et al, 2007;Habjan et al, 2008;Ianevski et al, 2018;Jupille et al, 2011;Kittel et al, 2004;Lee et al, 2017;Utt et al., 2016). In addition, qPCR/RT-qPCR, RNA/DNA sequencing, RNA/DNA hybridization, immuno-and plaque assays as well as CRISPR-CAS9 systems could be used for detection of inhibitory effects of BSAAs (Boonham et al, 2014;Fischer et al, 2017;Konig et al, 2019;Laamiri et al, 2018;Landry, 1990;Perez et al, 2013;Sashital, 2018;Zhou et al, 2018). Interestingly, CRISPR-Cas9, siRNA and shRNA approaches were used for identification of BSAA targets (Deans et al, 2016;Puschnik et al, 2017).…”

Discovery and Development of Safe-in-Man Broad-Spectrum Antiviral Agents

“…RFP-expressing RVFV, nanoLucexpressing CHIKV and RRV, as well as GFP-expressing FLUAV, HCV and HMPV also allowed identification of novel activities of several BSAAs (13,21,(25)(26)(27)(28)(29)(30)(31). In addition, qPCR/RT-qPCR, RNA/DNA sequencing, RNA/DNA hybridization, immuno-and plaque assays as well as CRISPR-CAS9 systems could be used for detection of inhibitory effects of BSAAs (32)(33)(34)(35)(36)(37)(38)(39). Interestingly, CRISPR-Cas9, siRNA and shRNA approaches were used for identification of BSAA targets (40,41).…”

Discovery and Development of Safe-in-Man Broad-Spectrum Antiviral Agents

“…RFPexpressing RVFV, nanoLuc-expressing CHIKV and RRV, as well as GFP-expressing FLUAV, HCV and HMPV also allowed identification of novel activities of several BSAAs (Andersen et al, 2019b;Bosl et al, 2019;de Graaf et al, 2007;Habjan et al, 2008;Ianevski et al, 2018;Jupille et al, 2011;Kittel et al, 2004;Lee et al, 2017;Utt et al, 2016). In addition, qPCR/RT-qPCR, RNA/DNA sequencing, RNA/DNA hybridization, immuno-and plaque assays as well as CRISPR-CAS9 systems could be used for detection of inhibitory effects of BSAAs (Boonham et al, 2014;Fischer et al, 2017;Konig et al, 2019;Laamiri et al, 2018;Landry, 1990;Perez et al, 2013;Sashital, 2018;Zhou et al, 2018). Interestingly, CRISPR-Cas9, siRNA and shRNA approaches were used for identification of BSAA targets (Deans et al, 2016;Puschnik et al, 2017).…”