Corporate Banner
Satellite Banner
Technology
Networks
Scientific Communities
 
Become a Member | Sign in
Home>News>This Article
  News
Return

CRISPR System Scales Up in Human Cells

Published: Friday, December 13, 2013
Last Updated: Friday, December 13, 2013
Bookmark and Share
For decades, researchers have sought a biological toolset capable of precisely and systematically turning off genes throughout the genomes of human cells.

The CRISPR-Cas9 system – a recently discovered system with bacterial origins – has the potential to overcome many of the limitations of currently available gene-silencing techniques. Earlier this year, several research groups showed that it was possible to use CRISPR-Cas9 to turn off genes in mammalian cells.

But in order to investigate and better understand the genetics of health and disease, scientists need a toolset that can reliably turn off each of the genes in the genome on a large scale.

In companion papers published together this week in Science, researchers from the Broad Institute, Whitehead Institute, McGovern Institute for Brain Research, and elsewhere have scaled up, demonstrating the capabilities of the CRISPR-Cas9 system in large-scale studies of several types of human cells, turning off genes accurately and efficiently at a genomic scale instead of just one or a few genes at a time. The two papers – which offer up libraries of tens of thousands of complexes tailored to match precise locations throughout the genome – lay the groundwork for a range of future studies, including investigations into neural development, cancer, and many other human diseases.

“We can now use this technology on a genome-wide scale, giving us the ability to interrogate any gene we want,” said Feng Zhang, a core member of the Broad Institute, an investigator at the McGovern Institute, and an assistant professor at MIT. Zhang is also the senior author of one of the Science papers. “Additionally, we demonstrate the utility of CRISPR for making biological discoveries, identifying new genes likely involved in how cancer cells become resistant to treatment. This is likely the first biological discovery made using CRISPR.”

Unlike other gene-silencing tools, the CRISPR-Cas9 system targets the genome’s source material: while RNA interference (RNAi) must target many copies of messenger RNA in order to tamp down a gene’s expression, CRISPR-Cas9 permanently turns off genes at the DNA level. The CRISPR-Cas9 system includes an enzyme that makes a cut in DNA, and is paired with a single guide RNA (sgRNA), which researchers construct to home in on a specific site in the genome. The DNA cut – known as a double strand break – closely mimics the kinds of mutations that occur naturally, for instance after chronic sun exposure. But unlike the UV rays that can result in genetic alterations, the CRISPR-Cas9 system causes a mutation at a precise location in the genome.

When cellular machinery repairs the DNA break, it removes a small snip of DNA, rendering a gene non-functional. In this way, researchers can precisely turn off specific genes in the genome.

To turn off genes on a grand scale, both sets of investigators developed libraries of more than 65,000 sgRNAs. The library from Zhang’s group is designed to target almost every protein-coding gene in the genome.

Both research teams used CRISPR-Cas9 to study the development of resistance to drugs used to treat cancer. The research team from the Whitehead and the Broad investigated genes whose loss conferred resistance to etoposide, a drug used to treat many forms of cancer including lung cancer and testicular cancer. The Broad and McGovern researchers used CRISPR-Cas9 to find genes involved in resistance to a drug commonly used to treat melanoma. In both sets of experiments, the research teams uncovered several previously unknown genes tied to resistance as well as many established genes.

In addition to these experiments on resistance, the researchers also conducted several other studies, using the CRISPR-Cas9 system in human pluripotent cells and cells grown in different culture conditions. The CRISPR-Cas9 system proved effective across these cell types, further demonstrating its versatility.

“With this work, it is now possible to conduct systematic genetic screens in mammalian cells,” said David Sabatini, a member of the Whitehead, professor of biology at MIT, investigator of the Howard Hughes Medical Institute, senior associate member at the Broad, and a member of the Koch Institute. Sabatini is also a co-senior author of one of the Science papers. “This will greatly aid efforts to understand the function of both protein-coding genes as well as non-coding genetic elements.”

The research teams compared the results of using the CRISPR-Cas9 system to turn off genes to the effectiveness of RNAi to turn down the signal of genes. In general, RNAi only partially reduces a gene’s signal – knocking it down – while CRISPR-Cas9 turns off the gene’s signal completely – knocking it out. RNAi is also prone to off-target effects – disrupting unintended gene targets – while CRISPR-Cas9 showed more consistent results.

David Root, director of the Broad’s Genetic Perturbation Platform (formerly known as the RNAi Platform), and his platform colleagues assisted in the development of the GeCKO (genome-scale CRISPR knockout) screening technique reported in the Broad-McGovern study. In addition to RNAi screening, the platform now offers a range of genetic perturbation technologies to the Broad community. That list now includes the CRISPR-Cas9 system.

“The CRISPR-Cas9 screening method distinguishes itself from RNAi by producing knockouts instead of knockdowns and it will be cleaner for many phenotypes to see the complete knockout,” said Root who is a co-author of the Broad-McGovern paper. “The agreement among the distinct reagents targeting the same gene looks a lot higher for CRISPR-Cas9 compared to RNAi, which gives you a lot more confidence about gene specificity for these results.”

One of the unique advantages to the CRISPR-Cas9 system that researchers intend to explore in the future is that the system offers access to the world beyond genes: non-coding regions of the genome that may influence when and where proteins are produced in a manner that does not depend on their production of RNA. Such regions were out of reach for RNAi, but with CRISPR-Cas9, these elements may now be exploitable in mammalian cells.

“These papers together demonstrate the extraordinary power and versatility of the CRISPR-Cas9 system as a tool for genome-wide discovery of the mechanisms underlying mammalian biology,” said Eric Lander, director of the Broad Institute and co-senior author of one of the Science papers. “And we are just at the beginning: we’re still uncovering the capabilities of this system and its many applications.”


Further Information
Access to this exclusive content is for Technology Networks Premium members only.

Join Technology Networks Premium for free access to:

  • Exclusive articles
  • Presentations from international conferences
  • Over 2,500+ scientific posters on ePosters
  • More Than 3,700+ scientific videos on LabTube
  • 35 community eNewsletters


Sign In



Forgotten your details? Click Here
If you are not a member you can join here

*Please note: By logging into TechnologyNetworks.com you agree to accept the use of cookies. To find out more about the cookies we use and how to delete them, see our privacy policy.

Related Content

Researchers Find Link Between Microbiome, Type 1 Diabetes
Largest longitudinal study to date sees shift in microbiome species diversity prior to onset of disease.
Friday, February 06, 2015
Scientific News
The Changing Tides of the In Vitro Diagnostics Market
With the increasing focus in personalized medicine, diagnostics plays a crucial role in patient monitoring.
LaVision BioTec Reports on the Neuro Research on the Human Brain After Trauma
Company reports on the work of Dr Ali Ertürk from the Institute for Stroke and Dementia Research at LMU Munich.
NIH Study Shows No Benefit of Omega-3 Supplements for Cognitive Decline
Research was published in the Journal of the American Medical Association.
Less May Be More in Slowing Cholera Epidemics
Mathematical model shows more cases may be prevented and more lives saved when using one dose of cholera vaccine instead of recommended two doses.
Investigating the Vape
Expert independent review concludes that e-cigarettes have potential to help smokers quit.
NIH Launches Human RSV Study
Study aims to understand infection in healthy adults to aid development of RSV medicines, vaccines.
Researchers Discover Synthesis of a New Nanomaterial
Interdisciplinary team creates biocomposite for first time using physiological conditions.
Poor Survival Rates in Leukemia Linked to Persistent Genetic Mutations
For patients with an often-deadly form of leukemia, new research suggests that lingering cancer-related mutations – detected after initial treatment with chemotherapy – are associated with an increased risk of relapse and poor survival.
Flu Remedies Help Combat E. coli Bacteria
Physiologists from the University of Zurich have now discovered why the intestinal bacterium Escherichia coli (E. coli) multiplies heavily and has an inflammatory effect.
Marijuana Genome Unraveled
A study by Canadian researchers is providing a clearer picture of the evolutionary history and genetic organization of cannabis, a step that could have agricultural, medical and legal implications for this valuable crop.
Scroll Up
Scroll Down
Skyscraper Banner

Skyscraper Banner
Go to LabTube
Go to eposters
 
Access to the latest scientific news
Exclusive articles
Upload and share your posters on ePosters
Latest presentations and webinars
View a library of 1,800+ scientific and medical posters
2,500+ scientific and medical posters
A library of 2,500+ scientific videos on LabTube
3,700+ scientific videos
Close
Premium CrownJOIN TECHNOLOGY NETWORKS PREMIUM FREE!