WARF: P130315US02

Gene Editing Tool Uses New Enzyme


James Thomson, Zhonggang Hou, Yan Zhang, Rakhi Rajan, Erik Sontheimer, Alfonso Mondragón

The Wisconsin Alumni Research Foundation (WARF) is seeking commercial partners interested in developing a method for DNA cleavage and genome editing of eukaryotic cells, including human stem cells.
OVERVIEWGenome editing has proven difficult in cells, especially mammalian cells. One way to improve efficiency is to introduce a double-strand break (DSB) in the desired DNA region. These DSBs stimulate cellular repair machinery and, in the presence of a suitable repair template, greatly speed up the DNA modification process.

Currently there are two main ways to introduce DSBs into the genomes of mammalian cells: zinc finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs). A major obstacle for both approaches is the difficulty of engineering peptides that recognize specific target DNA sites. Also, for each target sequence a different pair of ZFNs or TALENs must be engineered. The process is costly and time consuming.

Another method for genome editing uses CRISPRs (clustered regularly interspaced short palindromic repeats). Recently, CRISPR RNAs have been developed that direct DNA cleavage by a bacterial enzyme called Cas9. Several Cas9 proteins from Streptococcus species are believed to be effective, but each has its own targeting sequence requirements.

Maximizing the potential of CRISPR as a gene editing tool will require identifying alternative enzymes.
THE INVENTIONA UW–Madison researcher and collaborators have developed a method for RNA-directed DNA cleavage and gene editing using a new form of Cas9 enzyme. The new form, abbreviated NmCas9, comes from the Neisseria meningitides bacteria.

A CRISPR kit can be made containing RNA molecules that direct the new enzyme to bind to and cleave/nick a target sequence.
  • DNA cleavage and gene editing in eukaryotic cells, including human embryonic and induced pluripotent stem cells
  • User friendly
  • Target specific
  • Less likely to result in off-target effects
  • New enzyme can accommodate broader range of targeting RNA structures and functionalities.
  • Mutant forms of the enzyme can be made that bind to but do not cleave the DNA.
STAGE OF DEVELOPMENTThe researchers have demonstrated their method using human embryonic stem cells and induced pluripotent stem cells.
For More Information About the Inventors
Contact Information
For current licensing status, please contact Andy DeTienne at or 608-960-9857.
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