Naked-Eye Detection of Grapevine Red-Blotch Viral Infection Using a Plasmonic CRISPR Cas12a Assay

Abstract

Herein, we described a novel plasmonic CRISPR Cas12a assay for the visual, colorimetric detection of grapevine viral infections. Our assay generates rapid and specific colorimetric signals for nucleic acid amplicons by combining the unique target-induced incriminate single-stranded DNase activity of Cas12a with plasmon coupling of DNA functionalized gold nanoparticles. The practical applicability of our plasmonic assay was successfully demonstrated through the detection of emerging red-blotch viral infections in grapevine samples collected from commercial vineyards.

CRISPR-based DNA and RNA detection with liquid-liquid phase separation

“…Furthermore, DNA-functionalized gold nanoparticles can be used in various ways to generate bright signals after the target activation of Cas12a and its indiscriminate cleavage activity. This can be achieved through metal-enhanced fluorescence (40) or by inhibiting the assembly of DNA-functionalized plasmonic nanoparticles (41). The abovementioned assays outperform our method in terms of lower detection limits, which are 10 pM (ÀPCR), femtomolar (ÀPCR), and 10 aM (þPCR), respectively.…”

“…The readouts of nanoparticle-based detection methods are as simple as turbidity. They require either a ultraviolet light source (39) or the measurement of color changes (40,41). The production of nanoparticles, however, is arguably more complicated.…”

CRISPR-Cas based virus detection: Recent advances and perspectives

“…Utilization of this platform, African swine fever virus was detected by naked eye within 1 hour. Combining the trans -cleavage activity of Cas12a and DNA functionalized AuNPs, Li et al generated a plasmonic CRISPR-Cas12a assay for grapevine red blotch virus (GRBV) detection by naked eye ( Li et al, 2019d ). Hu et al developed a magnetic pull-down assisted colorimetric diagnosis based on the CRISPR-Cas12a system, termed as M-CDC ( Hu et al, 2020 ).…”

CRISPR‐Cas12a System for Biosensing and Gene Regulation

“…The observed absorption peak is shifted to shorter or longer wavelengths and simultaneously the color of the colloidal solution is changed between red and purple, and the test results can be clearly identified by the naked eye. The first colorimetric assay combined with CRISPR‐Cas12a involved the trans cleavage of ssDNA substrates as the linker of AuNPs to control the aggregation and dispersion behavior [26g,h] (Figure 3a). The ssDNA substrates of collateral cleavage can also be stabilizers preventing salt‐induced aggregation of bare AuNPs.…”