Research News
July 27, 2020
The Institute of Medical Science
The University of Tokyo
http://www.ims.u-tokyo.ac.jp/imsut/en/

Development of an accurate and rapid diagnostic
method for COVID-19 using CRISPR-Cas3―Paper test strip results are possible within 40 minutes―
A new diagnostic method for COVID-19, SARS-CoV-2, using the Japanese genome editing
technology CRISPR-Cas3 (*1) has been developed that can accurately detect COVID-19 in 40minutes. A research team working in The Institute of Medical Science, The University ofTokyo, published their method on the pre-print server medRxiv on June 2.
The new diagnostic method, named "CONAN (*2)," was the first to use CRISPR-Cas3 todetect the new coronavirus (SARS-CoV-2). The test is performed using a nasopharyngeal
swab to detect viral RNA.
CRISPR-Cas3 is a genome editing tool developed in Japan that artificially cuts DNA via amultiprotein complex. It was developed in 2019 by the research team leader, Professor
Tomoji Mashimo, The Institute of Medical Science, The University of Tokyo and otherresearchers.
CRISPR-Cas-3 is a Class 1 CRISPR system that can be programmed to target a defined
gene sequence. The team used this mechanism to detect trace amounts of viral RNA foundin genomic samples of patients infected with SARS-CoV-2.
The research team found a positive concordance rate (PPA) of 90% (9/10 cases) and anegative concordance rate (NPA) of 95.3% (20/21 cases), which rivals the PCR test method in
detection sensitivity and specificity. In addition, the team claimed success in detecting thepresence or absence of viral RNA down to the tens-of-copies level with a test strip in only 40
minutes.
Please see the medRxiv (https://www.medrxiv.org/content/10.1101/2020.06.02.20119875v1)
for more details.

Speed, simplicity, adaptability and low costTo summarize the results of this study, the CONAN method has the following advantages
over PCR and antigen test methods.
1) The virus can be detected within 40 minutes from patient-derived samples.
2) Only general reagents, test papers and constant temperature heat blocks are required fortesting.
3) The method can detect as few as several dozen viral RNA copies with high sensitivity andaccuracy.
4) Since CRISPR-Cas3 can detect even a single base difference, the method can be readily
adapted for detecting new circulating mutations in the virus found to confer drug resistanceor increased virulence.
5) The method is not limited to SARS-CoV-2; the nucleotide sequences of any virus can bedetected.
In a nutshell, by utilizing CONAN, the new COVID-19 diagnostic method can be said tocombine the advantages of PCR-based methods in sensitivity and accuracy with those of
antigen testing methods in speed, simplicity, and low cost (Figure 1).

Applicable to influenza diagnosisIn the future, the research team will make a kit through C4U (http://www.crispr4u.jp/en/), aJapanese university-based bio-venture company. The team intends to put the method into
practical use as a COVID-19 rapid diagnostic agent that can be easily used in the medical
field.
Professor Mashimo says: "If CONAN is put to practical use as a COVID-19 diagnostic
method and used in various places such as medical facilities, airports, and outdoors, it will
greatly contribute to the prevention of further spread of the new coronavirus."
According to the research team, CONAN also succeeded in detecting viral RNAs of type A(H1N1pdm09, H3N2), which account for more than 95% of influenza circulating every year,using the same method.
The research team has declared that they will proceed with the development of a new
influenza diagnostic at the same time.

2This project was supported in part by JSPS KAKENHI Grant Number 18H03974 (T.M.) and
19K16025 (K.Y.). Partially support also came from the Platform Project for Supporting Drug
Discovery and Life Science Research (Basis for Supporting Innovative Drug Discovery andLife Science Research (BINDS)) from AMED under Grant Number JP20am0101070(support numbers 1251 and 2463), from Research Program on Emerging and Re-emerging
Infectious Diseases from AMED (JP19fk0108113), and from the National Institutes ofAllergy and Infectious Diseases funded Center for Research on Influenza Pathogenesis(CRIP; HHSN272201400008C).

Figure 1 : Detection of new coronavirus RNA by lateral flow test paper (red arrow: positive)

Figure 2： Rapid diagnostic tests against infectious diseases

3Figure 3: CONAN method for the RNA virus detection＃＃＃Research Notes(*1) CRISPR-Cas3
Many bacteria have a defense system called "CRISPR-Cas system" that resembles adaptiveimmunity. CRISPR-Cas3 is a CRISPR system belonging to Class 1 among CRISPR systems ofbacteria and archaea. It was developed in 2019 as a domestic genome editing tool that artificiallycuts DNA with a multiprotein complex.
(*2) CONAN
Cas3 Operated Nucleic Acid detectioN method. By using a mixture of CRISPR-Cas3 protein thatrecognizes the target sequence and a DNA probe for detection as a biosensor, it is possible todetect quickly and accurately whether or not the target sequence is present in a sample.

About the research1) Article
Journal name: Pre-print server "medRxiv" (published on June 2)
Kazuto Yoshimi, Kohei Takeshita, Seiya Yamayoshi, Satomi Shibumura, Yuko Yamauchi, MasakiYamamoto, Hiroshi Yotsuyanagi, Yoshihiro Kawaoka, Tomoji Mashimo “Rapid and accurate
detection of novel coronavirus SARS-CoV-2 using CRISPR-Cas3” medRxiv : June 2.
DOI number: 10.1101/2020.06.02.20119875
URL: https://www.medrxiv.org/content/10.1101/2020.06.02.20119875v142) Contact

Research Contact
Professor Tomoji MashimoThe Institute of Medical Science, The University of Tokyo
https://www.ims.u-tokyo.ac.jp/imsut/en/index.html
Press Contact
Division for International Affairs office, The Institute of Medical Science, The University ofTokyo
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About IMSUT (The Institute of Medical Science, The University of Tokyo)
The Institute of Medical Science, The University of Tokyo (IMSUT) evolved from its origin,
the Institute for Infectious Disease in 1967. The mission of IMSUT is to advance basic
knowledge underlying infectious diseases, cancer and other intractable diseases andultimately to control them. IMSUT consists of about 165 faculty members, 224 graduate
students coming from various schools such as medicine, science, agriculture, pharmaceuticalscience, and engineering to develop more effective interdisciplinary research in basic life
science and genomic medicine.

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