Novel COVID-19 Test Uses CRISPR Technology

UCSF and Mammoth Biosciences use CRISPR to detect the SARS-CoV-2 coronavirus.

Posted Apr 17, 2020

Source: Piro4D/Pixabay

Testing for COVID-19 is a critical component in fighting the pandemic. The key is to have accurate and widespread testing. On April 16, 2020, scientists at the University of California San Francisco and Mammoth Biosciences published in Nature Biotechnology a study that uses CRISPR to detect SARS-CoV-2, the coronavirus that causes the COVID-19 disease, in less than an hour.  

The CRISPR-based diagnostic test for COVID-19 may offer certain advantages over both serology (blood serum) and real-time reverse transcription-polymerase chain reaction (RT-PCR) tests, according to the research team.

“Although serology tests are rapid and require minimal equipment, their utility may be limited for diagnosis of acute SARS-CoV-2 infection, because it can take several days to weeks following symptom onset for a patient to mount a detectable antibody response,” the researchers wrote in the study.

The researchers point out that while real-time reverse transcription-polymerase chain reaction (RT-PCR) tests can take around four to six hours, the typical turnaround time for screening and diagnosing patients for COVID-19 typically has been over 24 hours due to the need to ship the samples overnight to special laboratories. Furthermore, unlike RT-PCR tests, the new CRISPR-based test can be conducted with off-the-shelf reagents and common equipment found in nearly any lab.

CRISPR is an acronym that stands for Clustered Regularly Interspaced Short Palindromic Repeats. In biochemistry, CRISPR refers to a series of short repeating DNA sequences with spacers in between that is the distinctive characteristic of the genomes of prokaryotic organisms. Prokaryotes are single-celled organism such as bacteria and archaea that lacks a nucleus and membrane-bound organelles. In Greek, “pro” means before, and “karyo” refers to a kernel; hence, prokaryotes are aptly named. In contrast, animals, plants, fungi, and protists are eukaryotes that do have a membrane-bound nucleus and organelles.

CRISPR is a central part of the bacteria's immune system. The DNA of the virus is incorporated into the bacteria’s own genome by becoming the spacers, which confers the bacteria protection from the virus from future attacks.

In the field of genomics, biotechnology, and precision medicine, the term CRISPR refers to a specialized gene-editing technique, such as CRISPR-Cas9, where Cas is short for CRISPR-associated. When Cas genes located near CRISPR are activated, they produce Cas enzymes, which are proteins that have the ability to cut DNA.

For example, CRISPR-Cas9 has proven capable of editing human and animal DNA. In 2012, pioneering researchers Jennifer Doudna, Ph.D., at the University of California, Berkeley, and French scientist Emmanuelle Charpentier, Ph.D., at the Max Planck Institute for Infection Biology, discovered CRISPR-Cas9 capabilities and published a landmark study in Science with their research colleagues. In January 2013, scientist Feng Zhang, Ph.D., at the Broad Institute of MIT and Harvard, McGovern Institute for Brain Research at MIT, Massachusetts, along with his colleagues published in Science a research study on using the CRISPR-Cas9 system for genome engineering in human and mouse cells. In that same year, researchers from George Church’s lab at Harvard University published in Science a study using Cas9 for RNA-guided human genome engineering.

Fast-forward to April 2020, and researchers from UCSF and Mammoth Biosciences have developed a way to detect the SARS-CoV-2 coronavirus using a CRISPR-Cas12-based assay. Doudna is the Co-founder and Scientific Advisory Board Chair at Mammoth Biosciences, privately-held biotech founded in 2017 with headquarters in San Francisco and funding from Apple CEO Tim Cook, the Mayfield Fund, Brook Byers, Verily, Wireframe Ventures, Plum Alley, Decheng Capital, 8VC, and NFX, among other investors. Mammoth Biosciences’ advisory board members include Jeff Huber, the Co-founder of Grail and former Illumina board member, Lloyd Minor, the Dean at the Stanford University School of Medicine, and Maneesh Jain, the CEO and Co-founder at Mirvie.  

Mammoth Biosciences teamed with Dr. Charles Chiu at UCSF to configure the DETECTR platform to detect the coronavirus that causes COVID-19. The new test, called the “SARS-CoV-2 DETECTR” (DNA Endonuclease-Targeted CRISPR Trans Reporter), has not received formal approval for clinical use yet; however, it is undergoing clinical validation with the aim of accelerating the regulatory approval process through the U.S. Food and Drug Administration (FDA) Emergency Use Authorization (EUA).

The SARS-CoV-2 DETECTR examines sample RNA that is extracted from swabbing either the back of the nose and throat (nasopharyngeal) or the back of the throat through the mouth (oropharyngeal). The test conducts reverse transcription and isothermal amplification at the same time on the sample. Then Cas12 detection is performed. Cas12 guide RNAs (gRNAs) are programmed to target SARS-CoV-2 or related coronavirus strains. Specifically, the nucleoprotein (N) gene gRNA looks for SARS-CoV-2, and the envelope gene (E) gRNA looks for three coronavirus strains that are SARS-like. The researchers report that the test takes less than 40 minutes, which is significantly less time than RT-PCR tests.

“Our CRISPR-based DETECTR assay provides a visual and faster alternative to the U.S. Centers for Disease Control and Prevention SARS-CoV-2 real-time RT–PCR assay, with 95 percent positive predictive agreement and 100 percent negative predictive agreement,” reported the researchers in their study.

Through harnessing innovation across genomics, biochemistry, virology, and molecular biology, trail-blazing scientists have created a novel method to test for the SARS-CoV-2 coronavirus.

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