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Double Isothermal Amplification and CRISPR-Cas12a for Sensitive Detection of Citrinin

Zhang, Man LU ; Xue, Xiaoting LU ; Gong, Haiyue LU ; Liu, Baolin and Ye, Lei LU orcid (2021) In ACS Agricultural Science and Technology 1(10). p.1997-2005
Abstract
An analytical method is developed for ultrasensitive detection of citrinin using double isothermal amplification and CRISPR-Cas12a. Gold nanoparticles (AuNPs) modified with antigen and thiol-terminated, single-strand DNA (ssDNA) are used as a probe. The antigen-modified AuNPs compete with citrinin to bind to magnetic beads coated with an anticitrinin antibody. After a simple magnetic separation, the AuNPs are collected, and the ssDNA are released after they are washed with a dithiothreitol solution. The ssDNA is first amplified by an exponential amplification reaction and then used as a primer in a subsequent hybridization chain reaction to produce double-stranded DNA (dsDNA) that contains a protospacer adjacent motif to allow recognition... (More)
An analytical method is developed for ultrasensitive detection of citrinin using double isothermal amplification and CRISPR-Cas12a. Gold nanoparticles (AuNPs) modified with antigen and thiol-terminated, single-strand DNA (ssDNA) are used as a probe. The antigen-modified AuNPs compete with citrinin to bind to magnetic beads coated with an anticitrinin antibody. After a simple magnetic separation, the AuNPs are collected, and the ssDNA are released after they are washed with a dithiothreitol solution. The ssDNA is first amplified by an exponential amplification reaction and then used as a primer in a subsequent hybridization chain reaction to produce double-stranded DNA (dsDNA) that contains a protospacer adjacent motif to allow recognition by CRISPR-Cas12a. The dsDNA activates the Cas12a-gRNA to cleave a reporter ssDNA to generate a fluorescence signal. The developed analytical method has a low detection limit (0.127 ng mL–1) and a wide linear range (0.005–500 μg mL–1) for detection of citrinin. For detection of citrinin in oat and flour, recoveries of 97–104% and 105–111% are obtained, respectively. By combining double isothermal amplification with CRISPR-Cas12a, ultrahigh sensitivity and selectivity can be achieved for detection of toxins in food. (Less)
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author
; ; ; and
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
Ultrasensitive detection, Biobarcode, CRISPR-Cas12a, Exponential amplification reaction, Hybridization chain reaction, Citrinin
in
ACS Agricultural Science and Technology
volume
1
issue
10
pages
9 pages
publisher
The American Chemical Society (ACS)
external identifiers
  • scopus:85125573215
ISSN
2692-1952
DOI
10.1021/acsfoodscitech.1c00321
language
English
LU publication?
yes
id
52ef5489-9d96-4a06-a214-8e841b2673ec
date added to LUP
2022-09-29 09:26:28
date last changed
2023-02-15 09:08:43
@article{52ef5489-9d96-4a06-a214-8e841b2673ec,
  abstract     = {{An analytical method is developed for ultrasensitive detection of citrinin using double isothermal amplification and CRISPR-Cas12a. Gold nanoparticles (AuNPs) modified with antigen and thiol-terminated, single-strand DNA (ssDNA) are used as a probe. The antigen-modified AuNPs compete with citrinin to bind to magnetic beads coated with an anticitrinin antibody. After a simple magnetic separation, the AuNPs are collected, and the ssDNA are released after they are washed with a dithiothreitol solution. The ssDNA is first amplified by an exponential amplification reaction and then used as a primer in a subsequent hybridization chain reaction to produce double-stranded DNA (dsDNA) that contains a protospacer adjacent motif to allow recognition by CRISPR-Cas12a. The dsDNA activates the Cas12a-gRNA to cleave a reporter ssDNA to generate a fluorescence signal. The developed analytical method has a low detection limit (0.127 ng mL<sup>–1</sup>) and a wide linear range (0.005–500 μg mL<sup>–1</sup>) for detection of citrinin. For detection of citrinin in oat and flour, recoveries of 97–104% and 105–111% are obtained, respectively. By combining double isothermal amplification with CRISPR-Cas12a, ultrahigh sensitivity and selectivity can be achieved for detection of toxins in food.}},
  author       = {{Zhang, Man and Xue, Xiaoting and Gong, Haiyue and Liu, Baolin and Ye, Lei}},
  issn         = {{2692-1952}},
  keywords     = {{Ultrasensitive detection; Biobarcode; CRISPR-Cas12a; Exponential amplification reaction; Hybridization chain reaction; Citrinin}},
  language     = {{eng}},
  month        = {{10}},
  number       = {{10}},
  pages        = {{1997--2005}},
  publisher    = {{The American Chemical Society (ACS)}},
  series       = {{ACS Agricultural Science and Technology}},
  title        = {{Double Isothermal Amplification and CRISPR-Cas12a for Sensitive Detection of Citrinin}},
  url          = {{http://dx.doi.org/10.1021/acsfoodscitech.1c00321}},
  doi          = {{10.1021/acsfoodscitech.1c00321}},
  volume       = {{1}},
  year         = {{2021}},
}