RNA-Editing Tool a Fast, Easy Test for COVID-19 and Other Diseases

Rice University postdoctoral researcher Jie Yang led an effort to adapt Cas13 genome editing tools to serve as a highly sensitive detector for the presence of the SARS-CoV-2 virus, which causes COVID-19. Credit: Jeff Fitlow/Rice University

Cas13 Engineered To Simplify the Identification of Coronavirus

A new engineered CRISPR-based method accurately finds

The new platform was highly successful compared to PCR testing. In fact, it found 10 out of 11 positives and no false positives for the virus in tests on clinical samples directly from nasal swabs. The scientists showed their technique finds signs of SARS-CoV-2 in attomolar (10-18) concentrations.

The study will be published today (September 22, 2022) in the journal Nature Chemical Biology. It was led by chemical and biomolecular engineer Xue Sherry Gao at Rice’s George R. Brown School of Engineering and postdoctoral researchers Jie Yang of Rice and Yang Song of Connecticut.

Using structure-guided Cas13, researchers at Rice University and the University of Connecticut modified a gene editing tool to serve as a highly sensitive diagnostic test for the presence of the SARS-CoV-2 virus. They used an off-the-shelf electrochemical sensor to deliver results. Credit: Jie Yang/Rice University

Cas13, like its better-known cousin Cas9, is part of the system by which bacteria naturally defend themselves against invading phages. Since its discovery, CRISPR-Cas9 has been adapted by scientists to edit living

She said the key is a well-hidden, flexible hairpin loop near Cas13’s active site. “It’s in the middle of the protein near the catalytic site that determines Cas13’s activity,” Yang said. “Since Cas13 is large and dynamic, it was challenging to find a site to insert another functional domain.”

From left, Rice University undergraduate student Jeffrey Vanegas, chemical and biomolecular engineer Xue Sherry Gao and postdoctoral researcher Jie Yang led the effort to modify a gene editing tool to serve as a diagnostic test for the presence of the SARS-CoV-2 virus. Credit: Rice University

The research team fused seven different RNA binding domains to the loop, and two of the complexes were clearly superior. When they found their targets, the proteins would fluoresce, revealing the presence of the virus.

“We could see the increased activity was five- or six-fold over wild-type Cas13,” Yang said. “This number seems small, but it’s quite astonishing with a single step of protein engineering.

“But that was still not enough for detection, so we moved the whole assay from a fluorescence plate reader, which is quite large and not available in low-resource settings, to an electrochemical sensor, which has higher sensitivity and can be used for point-of-care diagnostics,” she said.

With the off-the-shelf sensor, Yang said the engineered protein was five orders of magnitude (100,000x) more sensitive in detecting the virus compared to the wild-type protein.

The lab wants to adapt its technology to paper strips like those in home COVID-19 antibody tests, but with much higher sensitivity and

“Different viruses have different sequences,” Yang said. “We can design guide RNA to target a specific sequence that we can then detect, which is the power of the CRISPR-Cas13 system.”

But because the project began just as the COVID-19 pandemic took hold, SARS-CoV-2 was a natural focus. “The technology is quite amenable to all the targets,” she said. “This makes it a very good option to detect all kinds of mutations or different coronaviruses.”

“We are very excited about this work as a combinational effort of structure biology, protein engineering, and biomedical device development,” Gao added. “I greatly appreciate all the efforts from my lab members and collaborators.”

Reference: “Structure-Guided Engineering of LwaCas13a with Enhanced Collateral Activity for Ultrasensitive Nucleic Acid Detection 22 September 2022, Nature Chemical Biology.
DOI: 10.1038/s41589-022-01135-y

Co-authors of the paper are Rice postdoctoral researcher Xiangyu Deng, undergraduate Jeffrey Vanegas, and graduate student Zheng You; graduate students Yuxuan Zhang and Zhengyan Weng of the University of Connecticut; microbiology supervisor Lori Avery and Kevin Dieckhaus, a professor of medicine, of UConn Health; Yi Zhang, an assistant professor of biomedical engineering at the University of Connecticut; and Yang Gao, an assistant professor of biosciences at Rice.

Xue Sherry Gao is the Ted N. Law Assistant Professor of Chemical and Biomolecular Engineering at Rice.

The National Science Foundation (2031242, 2103025), the Welch Foundation (C-1952, C-2033-20200401), and the Cancer Prevention and Research Institute of Texas (RR190046) supported the research.

Leave a Reply

This site uses Akismet to reduce spam. Learn how your comment data is processed.