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CRISPR/Cas14a-Based Isothermal Amplification for Profiling Plant MicroRNAs.
Yang, Hao; Chen, Junbo; Yang, Sen; Zhang, Ting; Xia, Xuhan; Zhang, Kaixiang; Deng, Sha; He, Guiping; Gao, Hong; He, Qiang; Deng, Ruijie.
Affiliation
  • Yang H; College of Biomass Science and Engineering, Healthy Food Evaluation Research Center, Sichuan University, Chengdu 610065, China.
  • Chen J; Analytical & Testing Center, Sichuan University, Chengdu, Sichuan 610064, China.
  • Yang S; College of Biomass Science and Engineering, Healthy Food Evaluation Research Center, Sichuan University, Chengdu 610065, China.
  • Zhang T; College of Biomass Science and Engineering, Healthy Food Evaluation Research Center, Sichuan University, Chengdu 610065, China.
  • Xia X; College of Biomass Science and Engineering, Healthy Food Evaluation Research Center, Sichuan University, Chengdu 610065, China.
  • Zhang K; School of Pharmaceutical Sciences, Key Laboratory of Targeting Therapy and Diagnosis for Critical Diseases, Zhengzhou University, Zhengzhou 450001, China.
  • Deng S; College of Biomass Science and Engineering, Healthy Food Evaluation Research Center, Sichuan University, Chengdu 610065, China.
  • He G; College of Biomass Science and Engineering, Healthy Food Evaluation Research Center, Sichuan University, Chengdu 610065, China.
  • Gao H; College of Biomass Science and Engineering, Healthy Food Evaluation Research Center, Sichuan University, Chengdu 610065, China.
  • He Q; College of Biomass Science and Engineering, Healthy Food Evaluation Research Center, Sichuan University, Chengdu 610065, China.
  • Deng R; College of Biomass Science and Engineering, Healthy Food Evaluation Research Center, Sichuan University, Chengdu 610065, China.
Anal Chem ; 93(37): 12602-12608, 2021 09 21.
Article in En | MEDLINE | ID: mdl-34506121
ABSTRACT
MicroRNAs (miRNAs) play key roles in biological processes in plants, such as stress resistance, yet can hardly be quantified by an enzyme-involved terminal polymerization process due to their 2'-O-methyl modifications at the 3'-terminal. Herein, we proposed a CRISPR/Cas14a-based rolling circle amplification (termed Cas14R) assay, allowing reverse transcription-free and demethylation-free detection of plant miRNAs with single-nucleotide resolution. The employment of target-templated rolling circle amplification circumvents the extension of the unaccessible 2'-O-methyl group at the 3'-terminal. Particularly, the activated Cas14a confers the trans-cleavage activity for identifying target single-stranded DNA sequences without the necessity of the protospacer adjacent motif, generalizing the detection of miRNA sequences and the integration of different isothermal amplification techniques. Ultimately, the Cas14R assay has been applied to profile miR156a to evaluate the ripeness process of banana, indicating its feasibility in analyzing the roles of miRNAs in biological processes of plants.
Subject(s)

Full text: 1 Collection: 01-internacional Database: MEDLINE Main subject: MicroRNAs Language: En Journal: Anal Chem Year: 2021 Document type: Article Affiliation country: China

Full text: 1 Collection: 01-internacional Database: MEDLINE Main subject: MicroRNAs Language: En Journal: Anal Chem Year: 2021 Document type: Article Affiliation country: China