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Photocontrolled crRNA activation enables robust CRISPR-Cas12a diagnostics.
Hu, Menglu; Qiu, Zhiqiang; Bi, Zirong; Tian, Tian; Jiang, Yongzhong; Zhou, Xiaoming.
  • Hu M; School of Life Sciences, South China Normal University, Guangzhou 510631, People's Republic of China.
  • Qiu Z; School of Life Sciences, South China Normal University, Guangzhou 510631, People's Republic of China.
  • Bi Z; Ministry of Education (MOE) Key Laboratory of Laser Life Science, College of Biophotonics, South China Normal University, Guangzhou 510631, People's Republic of China.
  • Tian T; Guangdong Provincial Key Laboratory of Laser Life Science, College of Biophotonics, South China Normal University, Guangzhou 510631, People's Republic of China.
  • Jiang Y; School of Life Sciences, South China Normal University, Guangzhou 510631, People's Republic of China.
  • Zhou X; School of Life Sciences, South China Normal University, Guangzhou 510631, People's Republic of China.
Proc Natl Acad Sci U S A ; 119(26): e2202034119, 2022 06 28.
Article in English | MEDLINE | ID: covidwho-1900769
ABSTRACT
CRISPR diagnostics based on nucleic acid amplification faces barriers to its commercial use, such as contamination risks and insufficient sensitivity. Here, we propose a robust solution involving optochemical control of CRISPR RNA (crRNA) activation in CRISPR detection. Based on this strategy, recombinase polymerase amplification (RPA) and CRISPR-Cas12a detection systems can be integrated into a completely closed test tube. crRNA can be designed to be temporarily inactivated so that RPA is not affected by Cas12a cleavage. After the RPA reaction is completed, the CRISPR-Cas12a detection system is activated under rapid light irradiation. This photocontrolled, fully closed CRISPR diagnostic system avoids contamination risks and exhibits a more than two orders of magnitude improvement in sensitivity compared with the conventional one-pot assay. This photocontrolled CRISPR method was applied to the clinical detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA, achieving detection sensitivity and specificity comparable to those of PCR. Furthermore, a compact and automatic photocontrolled CRISPR detection device was constructed.
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Full text: Available Collection: International databases Database: MEDLINE Main subject: Reagent Kits, Diagnostic / Bacterial Proteins / Reverse Transcriptase Polymerase Chain Reaction / Endodeoxyribonucleases / CRISPR-Associated Proteins / Clustered Regularly Interspaced Short Palindromic Repeats / CRISPR-Cas Systems Type of study: Diagnostic study / Prognostic study Limits: Humans Language: English Journal: Proc Natl Acad Sci U S A Year: 2022 Document Type: Article

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Full text: Available Collection: International databases Database: MEDLINE Main subject: Reagent Kits, Diagnostic / Bacterial Proteins / Reverse Transcriptase Polymerase Chain Reaction / Endodeoxyribonucleases / CRISPR-Associated Proteins / Clustered Regularly Interspaced Short Palindromic Repeats / CRISPR-Cas Systems Type of study: Diagnostic study / Prognostic study Limits: Humans Language: English Journal: Proc Natl Acad Sci U S A Year: 2022 Document Type: Article