Features | Partner Sites | Information | LinkXpress
Sign In
GLOBETECH PUBLISHING
GLOBETECH PUBLISHING
JIB

New Genome Engineering Technique Eases Gene Repair in Human Pluripotent Stem Cells

By BiotechDaily International staff writers
Posted on 29 Aug 2013
Image: RNA-Guided Nucleases (RGNs), based on naturally occurring Type II CRISPR-Cas systems, are programmable endonucleases that can be used to perform targeted genome editing (Photo courtesy of Addgene).
Image: RNA-Guided Nucleases (RGNs), based on naturally occurring Type II CRISPR-Cas systems, are programmable endonucleases that can be used to perform targeted genome editing (Photo courtesy of Addgene).
An RNA-guided, DNA-cleaving interference pathway from bacteria [the type II clustered, regularly interspaced, short palindromic repeats (CRISPR)-CRISPR-associated (Cas) pathway] has been adapted for use in eukaryotic cells, greatly facilitating genome editing.

The CRISPR/Cas (CRISPR associated) system is a new genome-engineering technology recently developed from prokaryotes' adaptive immune response systems. The CRISPR system uses a short, noncoding RNA (crRNA) to target a human codon-optimized Cas9 nuclease to complementary (protospacer) sequences in the host genome. These sequences or arrays are composed of direct repeats that are separated by similarly sized nonrepetitive spacers. CRISPR arrays, together with a group of associated proteins, confer resistance to phages, possibly by an RNA-interference-like mechanism.

Investigators at the University of Wisconsin (Madison USA) and their colleagues at Northwestern University (Evanston, IL, USA) used a CRISPR-Cas system identified in the bacterium Neisseria meningitidis, which is distinct from the commonly used Streptococcus pyogenes system, to demonstrate efficient genome engineering in human pluripotent stem cells (hPSCs).

Heretofore, only two CRISPR-Cas systems (from S. pyogenes and S. thermophilus), each with their own distinct targeting requirements and limitations, had been developed for genome editing. In addition, only limited information existed regarding homology-directed repair (HDR)-mediated gene targeting using long donor DNA templates in hPSCs with these systems.

The investigators wrote in the August 12, 2013, online edition of the journal Proceedings of the National Academy of Sciences of the United States of America (PNAS) that by employing a distinct CRISPR-Cas system from N. meningitidis, they were able to demonstrate efficient targeting of an endogenous gene in three hPSC lines using HDR. The Cas9 RNA-guided endonuclease from N. meningitidis (NmCas9) recognized a protospacer adjacent motif (PAM) different from those recognized by Cas9 proteins from S. pyogenes and S. thermophilus (SpCas9 and StCas9, respectively). Similar to SpCas9, NmCas9 was able to use a single-guide RNA (sgRNA) to direct its activity. Because of its distinct protospacer adjacent motif, the N. meningitidis CRISPR-Cas machinery increased the sequence contexts amenable to RNA-directed genome editing.

“Human pluripotent stem cells can proliferate indefinitely and they give rise to virtually all human cell types, making them invaluable for regenerative medicine, drug screening, and biomedical research,” said senior author Dr. James A. Thomson, professor of embryonic stem cell biology at the University of Wisconsin. “Our collaboration with the Northwestern team has taken us further toward realizing the full potential of these cells because we can now manipulate their genomes in a precise, efficient manner. With this system, there is the potential to repair any genetic defect, including those responsible for some forms of breast cancer, Parkinson’s, and other diseases.”

Related Links:
University of Wisconsin
Northwestern University



comments powered by Disqus

Channels

Drug Discovery

view channel

Ibuprofen May Restore Immune Function in Older Individuals

New research suggests that macrophages from the lungs of old mice respond differently to infections than those of young mice, and ibuprofen given to the mice reversed these changes. New research using lab mice suggests that the solution to more youthful immune function might already be a common over-the-counter pain reliever.... Read more

Therapeutics

view channel
Image: Hair follicle (blue) being attacked by T cells (green) (Photo courtesy of Christiano Lab/Columbia University Medical Center).

Hair Restoration Method Clones Patients’ Cells to Grow New Hair Follicles

Researchers have developed of a new hair restoration approach that uses a patient’s cells to grow new hair follicles. In addition, the [US] Food and Drugs Administration (FDA) recently approved a new drug... Read more

Lab Technologies

view channel
Image: Leica Microsystems launches the inverted research microscope platform Leica DMi8 (Photo courtesy of Leica Microsystems).

New Inverted Microscope Designed to Readily Adapt to Changing Research Demands

A new inverted microscope for biotech and other life science laboratories was designed to readily accommodate modifications and upgrades to allow it to keep current with changing research demands and interests.... Read more

Business

view channel

Partnership Established to Decode Bowel Disease

23andMe (Mountain View, CA,USA), a personal genetics company, is collaborating with Pfizer, Inc. (New York, NY, USA), in which the companies will seek to enroll 10,000 people with inflammatory bowel disease (IBD) in a research project designed to explore the genetic factors associated with the onset, progression, severity,... Read more
 
Copyright © 2000-2014 Globetech Media. All rights reserved.