Home' Technology Review : March April 2014 Contents 57
MIT TECHNOLOGY REVIEW
VOL.117 | NO.2
The biotechnology industry was born in
1973, when Herbert Boyer and Stanley
Cohen inserted foreign DNA that they
had manipulated in the lab into bacteria.
Within a few years, Boyer had cofounded
Genentech, and the company had begun
using E. coli modified with a human gene
to manufacture insulin for diabetics. In
1974, Jaenisch, then at the Salk Institute
for Biological Studies in San Diego, cre-
ated the first transgenic mouse by using
viruses to spike the animal’s genome with
a bit of DNA from another species. In
these and other early examples of genetic
engineering, however, researchers were
limited to techniques that inserted the for-
eign DNA into the cell at random. All they
could do was hope for the best.
It took more than two decades before
molecular biologists became adept at effi-
ciently changing specific genes in animal
genomes. Dana Carroll of the University
of Utah recognized that zinc finger nucle-
ases, engineered proteins reported by col-
leagues at Johns Hopkins University in
1996, could be used as a programmable
gene-targeting tool. One end of the protein
can be designed to recognize a particular
DNA sequence; the other end cuts DNA.
When a cell then naturally repairs those
cuts, it can patch its genome by copying
from supplied foreign DNA. While the
technology finally enabled scientists to
confidently make changes where they
want to on a chromosome, it’s difficult
to use. Every modification requires the
researcher to engineer a new protein tai-
lored to the targeted sequence—a difficult,
time-consuming task that, because the
proteins are finicky, doesn’t always work.
TALENs, another significant advance
in gene editing, came along in 2010.
TALENs are also proteins that find and
cut a desired DNA sequence—but tailoring
them to new gene targets is much easier.
While they represented a great improve-
ment over zinc fingers, however, TALENs
are large proteins that are cumbersome to
work with and deliver into cells.
CRISPR changed everything. It
replaces the DNA-targeting proteins
with a short bit of RNA that homes in on
desired genes. Unlike the complex pro-
teins, RNA—which has nearly the same
simple structure as DNA—can be made
routinely in the lab; a technician can
quickly synthesize the roughly 20-letter-
long sequences the method requires. The
system makes it easy for medical research-
ers to modify a genome by replacing,
deleting, or adding DNA.
CRISPR stands for “clustered regularly
interspaced short palindromic repeats”—
clusters of brief DNA sequences that read
similarly forward and backward, which
are found in many types of bacteria. Sci-
entists first observed the puzzling DNA
segments in the 1980s but didn’t under-
stand for almost two decades that they
are part of a bacterial defense system.
When a virus attacks, bacteria can incor-
porate sequences of viral DNA into their
own genetic material, sandwiching them
between the repetitive segments. The next
time the bacteria encounter that virus,
they use the DNA in these clusters to make
RNAs that recognize the matching viral
sequences. A protein attached to one of
these RNAs then cuts up the viral DNA.
Zinc finger nucleases
What is it?
A protein consisting of a DNA-
cutting enzyme and a DNA-grabbing
region that can be tailored to
recognize different genes.
Also a protein containing a DNA-
cutting enzyme and a DNA-grabbing
region that can be programmed to
recognize different genes, but it is
easier to design than zinc finger
A DNA-cutting protein guided by an
RNA molecule that is able to find the
specific gene of interest.
Pros and cons
It was the first programmable
genome-editing tool, but it relies
on proteins that can be difficult
to engineer for new gene targets.
Potentially dangerous off-target
cuts are also possible.
Though simpler and cheaper to
design than zinc finger nucleases,
TALEN proteins can still be difficult
to produce and deliver. Off-target
cuts are a problem.
This technique is affordable and
easy to use, and it works for high-
throughput, multi-gene experiments.
Like the other tools, it can make off-
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