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Field-Deployable Candidatus Liberibacter asiaticus Detection Using Recombinase Polymerase Amplification Combined with CRISPR-Cas12a
Published on: December 23, 2022
Detection of target DNA with a visual CRISPR-associated hyperbranched rolling circle amplification technique
Beibei Zhang1, Miao Li1, Yuanmeng Wei1
1Henan Provincial People's Hospital, People's Hospital of Zhengzhou University, People's Hospital of Henan University, Henan Eye Institute, Henan Eye Hospital and Henan Key Laboratory of Ophthalmology and Visual Science, Zhengzhou, China.
Abstract:
This paper presents a novel clustered regularly interspaced short palindromic repeat (CRISPR)-associated HRCA technique (CART). During the entire detection process of CART, the target DNA is first specifically recognized and cleaved by a pair of Cas9/sgRNA complexes; then, the cleaved product is ligated into circular DNA as the template of HRCA, and the circular DNA is efficiently amplified by HRCA. Therefore, CART has the advantages of Cas9/sgRNA (single-base mismatch specificity) and HRCA (isothermal reaction temperature and high sensitivity). This technique has been verified by detecting various human papillomavirus (HPV) genes with numerous subtypes. In summary, this study provides a new and effective method for the detection of nucleic acids.
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