CRISPR/Cas9
Brian Tabone
What is CRISPR/Cas9?
• The CRISPR/CAS9 system is comprised of clustered regularlyinterspersed short palindromic repeated (CRISPR) DNA sequences
and CRISPR-associated genes (Cas) which code for proteins which
can read these sequences and locate them in other DNA strands and
perform operations on them (such as cut out or split and inject new
sequences).
• The CRISPR/Cas system evolved in nature as an immune defense
mechanism against viruses which attack bacteria. In nature, the
system warehouses DNA snippets from previous invaders and uses
this library to quickly identify and neutralize invading virus DNA
when it is next encountered.
• CRISPR/Cas9 has been adapted by science for precise, repeatable,
low cost gene editing that works in a multitude of species including
in humans.
CRISPR/Cas Immune Function
CRISPR/Cas9 history
• Scientists discovered the CRISPR/CAS system in the early 80s when
studying strains of the E. coli bacteria, but where not certain of its
function.
• The immune function of this system was confirmed in 2007 by R.
Barrangou and colleagues during their studies of immune response
to bacteriophage viruses attacking S. thermophilus bacteria (a
bacteria common in yogurt).
• In 2012, Doudna and Charpentier labs discovered the potential for
reliable, easy gene editing through application of the CRISPR/Cas9
version of the CRISPR/Cas system
What makes CRISPR/Cas9 valuable
• Previous gene editing techniques such as Zinc-finger nuclease
enhanced gene targeting and TALENs are more costly, difficult, and
error prone.
• New England BioLabs reports: “in human cells, custom-designed
ZFNs and TALENs could only achieve efficiencies ranging from 1% to
50%. In contrast, the Cas9 system has been reported to have
efficiencies up to >70% in zebrafish and plants, and ranging from 2–
5% in induced pluripotent stem cells.”
• They type II CRISPR nuclease simplicity and the requirements of
only 3 components (Cas9, crRNA and trRNA) make this system easy
to adapt for gene editing.
• CRISPR/Cas9 promises to lower the cost, improve accuracy, and
increase availability of genetic engineering to a broader spectrum of
scientists and engineers
CRISPR/Cas9 Gene Editing
• Previous gene editing techniques such as Zinc-finger nuclease
enhanced gene targeting and TALENs are more costly, difficult, and
error prone.
• New England BioLabs reports: “in human cells, custom-designed
ZFNs and TALENs could only achieve efficiencies ranging from 1% to
50%. In contrast, the Cas9 system has been reported to have
efficiencies up to >70% in zebrafish and plants, and ranging from 2–
5% in induced pluripotent stem cells.”
• They type II CRISPR nuclease simplicity and the requirements of
only 3 components (Cas9, crRNA and trRNA) make this system easy
to adapt for gene editing.
• CRISPR/Cas9 promises to lower the cost, improve accuracy, and
increase availability of genetic engineering to a broader spectrum of
scientists and engineers
What can we do with CRISPR/Cas9
• Control gene expression by activating or repressing target
sequences
• Identify sequence locations in a genome using fluorescent
molecules on nondestructive Cas9 enzymes
• Treat Cystic Fibrosis by inserting the missing gene sequence.
Correct the missing sequence so that is not passed down through
subsequent generations.
• Edit the gene sequence of mosquitos to either reduce their
reproduction rate or reduce their ability to carry diseases like
Malaria.
• Rapidly develop transgenic models which expands biological
research beyond traditional animal model organisms.
Risks
• CRISPR/Cas9 may make it easier for bad actors to create
weaponized diseases.
• A black market for gene editing of babies may become possible
due to the low cost and ready availability of the technology coupled
with the desire for improved competitiveness of one's children.
• Make it easier for corporations to make genetically modified
organisms for commercial use, with all the unintended
consequences that implies.
References
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Genome Editing Overview
New England BioLabs
Cell.com
YouTube video on Gene Editing with CRISPR/Cas9
CRISPR's Most Exciting Uses Have Nothing to Do With Gene-Editing
Scientists Call for Caution in Use of CRISPR/Cas9 Technology