2. Academic Validation
  3. CRISPR-Cas13a mediated nanosystem for attomolar detection of canine parvovirus type 2

CRISPR-Cas13a mediated nanosystem for attomolar detection of canine parvovirus type 2

  • Chin Chem Lett. 2019 Dec;30(12):2201-2204. doi: 10.1016/j.cclet.2019.10.032.
Haroon Khan 1,  Adeel Khan 1,  Yufeng Liu 2 3,  Su Wang 2 3,  Sumaira Bibi 4,  Hongpan Xu 2,  Yuan Liu 1,  Samran Durrani 5,  Lian Jin 6,  Nongyue He 1 6 7,  Tao Xiong 3
Affiliations

Affiliations

  • 1 State Key Laboratory of Bioelectronics, School of Biological Science and Medical Engineering, Southeast University, Nanjing 210096, China.
  • 2 Department of Clinical Laboratory, The Affiliated Drum Tower Hospital of Nanjing University Medical School, Nanjing 210008, China.
  • 3 College of Life Science, Yangtze University, Jingzhou 434025, China.
  • 4 Bio Resources Conservation Institute, National Agriculture Research Centre, Islamabad 350000, Pakistan.
  • 5 Laboratory of Biointerface & Biomaterials, School of Biological Science and Medical Engineering, Southeast University, Nanjing 210096, China.
  • 6 Hunan Key Laboratory of Biomedical Nanomaterials and Devices, Hunan University of Technology, Zhuzhou 412007, China.
  • 7 National Center for International Bio-targeting Theranostics, Guangxi Key Laboratory of Bio-targeting Theranostics, Collaborative Innovation Center for Targeting Tumor Theranostics, Guangxi Medical University, Nanning 530021, China.
PMID: 32288403 DOI: 10.1016/j.cclet.2019.10.032
Abstract

Canine parvovirus type 2 (CPV-2) Infection is the most lethal disease of dogs with higher mortality in puppies worldwide. In today's world, dogs are an integral part of our communities as well as dogs breeding and rearing has become a lucrative business. Therefore, a fast, accurate, portable, and cost-effective CPV-2 detection method with the ability for on-site detection is highly desired. In this study, we for the first time proposed a nanosystem for CPV-2 DNA detection with RNA-guided RNA endonuclease Cas13a, which upon activation results in collateral RNA degradation. We expressed LwCas13a in prokaryotic expression system and purified it through nickel column. Activity of Cas13a was verified by RNA-bound fluorescent group while using a quenched fluorescent probe as signals. Further Cas13a was combined with Recombinase polymerase amplification (RPA) and T7 transcription to establish molecular detection system termed specific high-sensitivity enzymatic reporter un-locking (SHERLOCK) for sensitive detection of CPV-2 DNA. This nanosystem can detect 100 amol/L CPV-2 DNA within 30 min. The proposed nanosystem exhibited high specificity when tested for CPV-2 and other dog viruses. This CRISPR-Cas13a mediated sensitive detection approach can be of formidable advantage during CPV-2 outbreaks because it is time-efficient, less laborious and does not involve the use of sophisticated instruments.

Keywords

Attomolar detection; CPV; CRISPR-Cas13a; RPA; SHERLOCK.

Figures
Products