4.7 Article

An all-in-one telomerase assay based on CRISPR-Cas12a trans-cleavage while telomere synthesis

Journal

ANALYTICA CHIMICA ACTA
Volume 1159, Issue -, Pages -

Publisher

ELSEVIER
DOI: 10.1016/j.aca.2021.338404

Keywords

Telomerase activity; CRISPR-Cas12a; Trans-cleavage; All-in-one; Point-of-care diagnosis

Funding

  1. National Natural Science Foundation of China [81873972, 81873980]
  2. Chongqing Science Fund for Distinguished Young Scholars [cstc2019jcyjjqX0 028]
  3. Training Program for Advanced Young Medical Personnel of Chongqing [2017HBRC003]
  4. Foundation for Innovative Research Groups of Chongqing Higher Education Institutions [CXQT20013]

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This new telomerase activity detection method based on CRISPR technology is fast, sensitive, and has a wide linear regression range, offering great potential for cancer diagnosis in a one-step process.
As one of the crucial factors associated with human life span and cancer progression, telomerase is regarded as an emerging biomarker for cancer diagnosis. Therefore, a facile, rapid and sensitive approach for telomerase activity detection with point-of-care (POC) diagnosis potential is in great demands. Herein, an all-in-one telomerase activity detection assay was established based on the telomere synthesis activated CRISPR-Cas12a system. A telomerase extension reaction generated telomere repeats sequences (TTAGGG)(n), which was recognized by a customized CRISPR-guided RNA (crRNA) simultaneously, and finally activated a typical trans-cleavage based CRISPR-Cas12a detection assay. With the inherent sensitivity of CRISPR-Cas12a, this approach achieved a great linear regression ranging from 100 to 2000 HeLa cells and a limitation of detection down to 26 HeLa cells. Moreover, by using the proposed method, telomerase can be detected in one pot under isothermal condition (37 degrees C) by a simple and fast workflow (one step within 1 h). Due to its excellent performance, this all-in-one method shows great potential in POC detection of the telomerase activity. (C) 2021 Elsevier B.V. All rights reserved.

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