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author:

Wang, Liu (Wang, Liu.) [1] | He, Fang (He, Fang.) [2] | Chen, Xueyun (Chen, Xueyun.) [3] | He, Kaiyu (He, Kaiyu.) [4] | Bai, Linlin (Bai, Linlin.) [5] | Wang, Qiang (Wang, Qiang.) [6] | Zhang, Fang (Zhang, Fang.) [7] (Scholars:张芳) | Xu, Xiahong (Xu, Xiahong.) [8]

Indexed by:

EI Scopus SCIE

Abstract:

CRISPR/Cas12a, as a powerful and programmable biosensing tool, has brought great convenience in visual detection of the target in a sequence-specific mode. Though fluorescent output is the predominant way of present visual methods, this strategy highly depends on cleaving fluorophore-/quencher-labeled oligonucleotides. Few research focuses on realizing fluorescent visual detection without DNA-labeling. Herein, we have proposed a CRISPR/Cas12a-based label-free fluorescent visual detection strategy. The basic principle of this strategy is that G-quadruplex can enhance the fluorescent emission of fluorescent ligands while dsDNA target-activated Cas12a make them impotent by disrupting the higher-ordered structure. After comparing four kinds of G-quadruplexspecific ligands, we selected Thioflavin T (ThT) to achieve the visual observation goal. The application feasibility was confirmed by combining with loop-mediated isothermal amplification (LAMP) for detection of Vibrio parahaemolyticus (V. parahaemolyticus), and it demonstrated a high sensitivity (1.36 x 10(2) copies) and specificity (among 12 kinds of bacteria). The method displayed similar performance as the real-time PCR for detection of real samples yet had lower requirement on the instrument. The signal output format could also distinguish Salmo salar from other seven kinds of sequence-similar Salmonidae. The findings and strategy proposed in this manuscript will expand the application of CRISPR/Cas12a-based label-free fluorescent analysis.

Keyword:

CRISPR/Cas12a G-quadruplex Label-free Thioflavin-T Vibrio parahaemolyticus Visual detection

Community:

  • [ 1 ] [Wang, Liu]Zhejiang Acad Agr Sci, Inst Agroprod Safety & Nutr, State Key Lab Managing Biot & Chem Threats Qual &, Hangzhou 310021, Peoples R China
  • [ 2 ] [He, Fang]Zhejiang Acad Agr Sci, Inst Agroprod Safety & Nutr, State Key Lab Managing Biot & Chem Threats Qual &, Hangzhou 310021, Peoples R China
  • [ 3 ] [Chen, Xueyun]Zhejiang Acad Agr Sci, Inst Agroprod Safety & Nutr, State Key Lab Managing Biot & Chem Threats Qual &, Hangzhou 310021, Peoples R China
  • [ 4 ] [He, Kaiyu]Zhejiang Acad Agr Sci, Inst Agroprod Safety & Nutr, State Key Lab Managing Biot & Chem Threats Qual &, Hangzhou 310021, Peoples R China
  • [ 5 ] [Bai, Linlin]Zhejiang Acad Agr Sci, Inst Agroprod Safety & Nutr, State Key Lab Managing Biot & Chem Threats Qual &, Hangzhou 310021, Peoples R China
  • [ 6 ] [Wang, Qiang]Zhejiang Acad Agr Sci, Inst Agroprod Safety & Nutr, State Key Lab Managing Biot & Chem Threats Qual &, Hangzhou 310021, Peoples R China
  • [ 7 ] [Xu, Xiahong]Zhejiang Acad Agr Sci, Inst Agroprod Safety & Nutr, State Key Lab Managing Biot & Chem Threats Qual &, Hangzhou 310021, Peoples R China
  • [ 8 ] [Chen, Xueyun]Fuzhou Univ, Coll Biol Sci & Engn, Fuzhou 350108, Peoples R China
  • [ 9 ] [Bai, Linlin]Fuzhou Univ, Coll Biol Sci & Engn, Fuzhou 350108, Peoples R China
  • [ 10 ] [Zhang, Fang]Fuzhou Univ, Coll Biol Sci & Engn, Fuzhou 350108, Peoples R China
  • [ 11 ] [Wang, Liu]Minist Agr & Rural Affairs, Key Lab Informat Traceabil Agr Prod, Hangzhou 310021, Peoples R China
  • [ 12 ] [He, Kaiyu]Minist Agr & Rural Affairs, Key Lab Informat Traceabil Agr Prod, Hangzhou 310021, Peoples R China
  • [ 13 ] [Xu, Xiahong]Minist Agr & Rural Affairs, Key Lab Informat Traceabil Agr Prod, Hangzhou 310021, Peoples R China
  • [ 14 ] [He, Fang]Ningbo Univ, Coll Food & Pharmaceut Sci, Ningbo 315800, Peoples R China

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Source :

SENSORS AND ACTUATORS B-CHEMICAL

ISSN: 0925-4005

Year: 2022

Volume: 370

8 . 4

JCR@2022

8 . 0 0 0

JCR@2023

ESI Discipline: CHEMISTRY;

ESI HC Threshold:74

JCR Journal Grade:1

CAS Journal Grade:1

Cited Count:

WoS CC Cited Count: 6

SCOPUS Cited Count: 7

ESI Highly Cited Papers on the List: 0 Unfold All

WanFang Cited Count:

Chinese Cited Count:

30 Days PV: 3

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