Journal of Animal Science and Technology
Korean Society of Animal Science and Technology

Development of a nucleic acid detection method based on the CRISPR-Cas13 for point-of-care testing of bovine viral diarrhea virus-1b.

Sungeun Hwang1, Wonhee Lee2, Yoonseok Lee1,3,*
1School of Biotechnology, Hankyong National University, Anseong 17579, Korea.
2Biattic Inc., Anyang 14059, Korea.
3Center for Genetic Information, Hankyong National University, Anseong 17579, Korea.
*Corresponding Author: Yoonseok Lee, School of Biotechnology, Hankyong National University, Anseong 17579, Korea, Republic of. Center for Genetic Information, Hankyong National University, Anseong 17579, Korea, Republic of. E-mail:

© Copyright 2023 Korean Society of Animal Science and Technology. This is an Open-Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License ( which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Received: Mar 17, 2023; Revised: Jul 17, 2023; Accepted: Jul 24, 2023

Published Online: Jul 25, 2023


Bovine Viral Diarrhea (BVD) is a single-stranded, positive-sense ribonucleic acid (RNA) virus belonging to the genus Pestivirus of the Flaviviridae family. BVD frequently causes economic losses to farmers. Among bovine viral diarrhea virus (BVDV) strains, BVDV-1b is predominant and widespread in Hanwoo calves. Reverse-transcription polymerase chain reaction (RT-PCR) is an essential method for diagnosing BVDV-1b and has become the gold standard for diagnosis in the Republic of Korea. However, this diagnostic method is time-consuming and requires expensive equipment. Therefore, Clustered Regularly Interspaced Short Palindromic Repeats-Cas (CRISPR-Cas) systems have been used for point of care (POC) testing of viruses. Developing a sensitive and specific method for POC testing of BVDV-1b would be advantageous for controlling the spread of infection. Thus, this study aimed to develop a novel nucleic acid detection method using the CRISPR-Cas13 system for POC testing of BVDV-1b. The sequence of the BVD virus was extracted from NCBI (NC_001461.1), and the 5' UTR region, commonly used for detection, was selected. CRISPR RNA (crRNA) was designed using the Cas13 design program and optimized for the expression and purification of the LwCas13a protein. Madin Darby bovine kidney (MDBK) cells were infected with BVDV-1b, incubated, and the viral RNA was extracted. To enable POC viral detection, the compatibility of the CRISPR-Cas13 system was verified with a paper-based strip through collateral cleavage activity. Finally, a colorimetric assay was used to evaluate the detection of BVDV-1b by combining the previously obtained crRNA and Cas13a protein on a paper strip. In conclusion, the CRISPR-Cas13 system is highly sensitive, specific, and capable of nucleic acid detection, making it an optimal system for the early point-of-care testing of BVDV-1b.

Keywords: BVDV-1b; CRISPR-Cas13 system; Point-of-care testing; Korean native cattle; nucleonic acid detection