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Biotechnology News

New Biotech Makes It Much Easier to Genetically Modify

Monkeys
A new gene-editing technique could lead to more useful animal models of disease, and perhaps one day more effective gene therapy for humans
Jan 31, 2014 | By Christine Gorman |

Like many babies, the wide-eyed twins are cute. The fact that they are macaque
monkeys is almost beside the point. What is not beside the point, however, is their
genetic heritage. These baby macaques are, as reported in Cell, the first primates to
have been genetically modified using an extremely precise gene-editing tool based on
the so-called CRISPR/Cas system.
Conducted by researchers in China, the new study is significant because it paves the
way for the custom development of laboratory monkeys with genetic profiles that are
similar to those found in humans with certain medical disorders. Although mice and
rats have long been the animals of choice when creating living models of human
disease, they have not been very helpful for studying neurological conditions such as
autism and Alzheimers disease; the differences between rodent and human brains
are just too great.

Genetically modified long-tailed macaques.

Credit: Cell, Niu et al.

To be sure, a few other genetically modified monkeys have been born over the past
decade and a half, but the methods used to alter their DNA were not as efficient or as
easy to use as the CRISPR/Cas technology. The amount of genome engineering in
monkeys is pretty small, says George Church, a professor of genetics at Harvard
Medical School. So yes, this [paper] is a pretty big deal.

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CRISPR stands for clustered regularly interspaced short palindromic repeats and refers to what at first glance appear to be meaningless
variations and repeats in the sequence of molecular letters (A, T, C and G) that make up DNA. These CRISPR patterns are found in
many bacteria and most archaea (an ancient group of bacteria that is now considered to be different enough from other one-celled
organisms to merit is own taxonomic kingdom, along with bacteria, protists, fungi, plants and animals).
First identified in bacteria in 1987, CRISPR elements started being widely used to create genetic engineering tools only in 2013. It took
that long to figure out that the patterns actually served a purpose, determine out what that purpose washelping archaea and bacteria
to recognize and defend themselves against virusesand then adapt that original function to a new goal.
Basically, biologists learned that certain proteins associated with the CRISPR system (dubbed, straightforwardly enough, CRISPRassociated, or Cas, proteins) act like scissors that cut any strands of DNA they come across. These cutting proteins, in turn, are guided to
specific strands of DNA by complementary pieces of RNA (a sister molecule to DNA). The bacteria generate specific guide strands of
RNA whenever they encounter a virus that is starting to hijack their cellular machinery. The guide-RNA complements the viral DNA,
which is how the Cas proteins know where to cut. The bacteria then keep a copy of the viral DNA in their own genetic sequence between
two CRISPR elements for future reference in case a similar virus tries to cause trouble later on.
In the past couple of years researchers have learned how to trick the Cas proteins into targeting and slicing through a sequence of DNA

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New Biotech Makes It Much Easier to Genetically Modify Mo...

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of their own choosing. By developing strands of RNA that precisely complement the part of the DNA molecule that they want to change,
investigators can steer the Cas proteins to a predesignated spot and cut out enough genetic material to permanently disrupt the usual
expression of the DNA molecule at that location.
In essence, scientists have turned a bacterial self-defense mechanism into an incredibly precise gene-editing tool. By some accounts
CRISPR technology has been successfully tried out on 20 different kinds of higher organisms (meaning higher than bacteria) in just the
past year or so.
The authors of the Cell study wrote that they found it relatively easy to adapt the CRISPR technology to monkey embryos. First they
introduced slight genetic changes targeting three genes on 22 fertilized macaque eggs. (The changes were not designed to create a
specific disease model in macaques, just to test the technique.) The manipulation produced 15 normally developing embryosof which
all but one showed evidence of the desired genetic changes. Satisfied with this initial success rate, the researchers then expanded their
efforts with the intention of producing a few fully developed baby monkeys.
The scientists collected 198 macaque eggs and injected them with macaque sperm. They then managed to further inject 186 fertilized
eggs with customized strips of RNA that generated and activated their own Cas proteins. These proteins in turn sliced the DNA double
helix at the precise location to which the guide RNA strands had directed them. The researchers then implanted 83 of the resulting
viable fertilized eggs into 29 macaque females. Ten of the monkeys became pregnant: three of them each carried a set of twins; three
carried triplets; and the remaining four carried single fetuses. So far, a set of twins were delivered by cesarean section after a normal
gestation period of 155 days.
The main purpose of the Cell study was to prove that CRISPR technology can be made to work in macaque monkeysan especially
important goal because macaques have become the new stand-in for people now that chimpanzees are being used less frequently for
medical research. The immediate next steps, says study co-author Xingxu Huang of Nanjing University is to try to make the genetic
modification process more efficient so a greater percentage of the fertilized eggs are appropriately modified and become viable animals.
And what about people? We believe the success of this strategy in nonhuman primates gives lots of potential for its application in
humans, Huang wrote in an e-mail. But we think, due to safety issues, it will take a long [time to expand] this strategy to human
embryos.

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