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DNA is the kind of polymer that chemists dream about. Because its complementary sequences can bind to one another, individual molecules of the right sequence will assemble all by themselves into intricate shapes and structures at the nanoscale. DNA can weave together and bind other molecules, allowing it to serve as a scaffold for complex nanomachinery.

DNA nanoengineering is dreamy, but difficult. Researchers have been putting together carefully chosen segments of DNA to form sheets, tubes, even simple machines such as tweezers since the early 1980s. But back then, designing these structures could take months to years. And because researchers were focused on designing them from scratch, they could use only the short segments, no more than 150-base-pairs long, that DNA synthesizers could manufacture. This in turn constrained the size and complexity of the designs. "The problem is that we don't just want to make small stuff, we want to make complicated small stuff, cheaply and easily," says Paul Rothemund, a computational bioengineer at the California Institute of Technology in Pasadena.

Rothemund wondered whether he could create the complicated stuff using a longer, naturally occurring piece of DNA, such as the genome of a virus, and folding it over on itself. So in 2004 and 2005 he spent months, he says, programming in his underpants, trying to work out a way to bend a 7,000-base-pair viral genome to his will. In his design he visualized how the genome could be folded into a predetermined, two-dimensional shape. Knowing the sequence of the virus at every twist and turn, he was able to write complementary DNA sequences, about 16-base-pairs long, that would essentially staple the folds in place. He ordered the 'staples' from a DNA-synthesis company, mixed them with his virus in a buffer that stabilized the DNA and then heated and cooled the mixture, allowing the single stranded viral DNA to bind with the staples (see 'Stapling a smiley'). The result, viewed using atomic-force microscopy, was the smiley face and several other shapes, created by what he called DNA origami1.

Credit: P. W. K. ROTHEMUND

The ease of DNA origami was a breakthrough, dispensing with the intricacies of precise DNA engineering and other metamaterials development. "It's like being able to bake a cake and not pay attention to the ingredient ratios," says Rothemund. But with the right ingredients complex structures can be built with the kind of precision that many people have been looking for. Origami scaffolds, sheets or bricks of folded DNA, are packed with known sequences that could be used to position DNA-binding molecules just a few nanometres apart. And the new, larger structures can contain upwards of 200 sites for affixing such molecules, compared with only a handful on pre-origami structures. This type of precision engineering could be a boon to nanoengineers wanting to position components on nanoelectronic circuits or for bioengineers looking to place proteins in close, accurate proximity to one another.

Now the challenge is to go beyond the novelty of Rothemund's smileys and a dozen or so other demonstration patterns and build structures with a practical purpose. Here's what several researchers are dreaming of doing.

Make a ruler

Rothemund's technique was a door opener for Friedrich Simmel, a biophysicist at the Technical University of Munich in Germany. Suddenly, Simmel says, he was able to have even "rather sloppy" physics students making DNA structures with ease. Simmel has used DNA origami to make a ruler to measure distances between single molecules and calibrate super-high resolution microscopes.

Simmel designed a DNA origami rectangle measuring 100 nm by 70 nm and included some staple strands labelled with fluorescent dye molecules. When the DNA folds, two labelled staples sit at opposite ends of the rectangle in precise locations. This ruler can be used to calibrate high-tech microscopes that can resolve objects smaller than the diffraction limit of light — roughly 200 nm (ref. 2). The kinds of molecules generally used for calibrating such scopes, such as loose pieces of DNA or filamentous proteins, are not ideal because they are flexible and their dimensions can change.

Simmel wants to use the ruler and related structures to help track the movement of molecular motors. Because fluorescent tags allow them to use light microscopy, as opposed to atomic-force microscopy or electron microscopy, researchers will be able to view molecular processes as they happen. That's important, Rothemund says, once scientists begin coupling proteins to DNA origami. Rulers like Simmel's will be useful to watch how those proteins behave.

Build an artificial leaf

DNA origami could allow researchers to put biomolecules together according to specification. A grail of sorts for many engineers working at the nanoscale is photosystem II, a complex of more than 20 protein subunits and accessories that helps to split water into hydrogen ions and oxygen during photosynthesis.

Attempts to recreate photosystem II, or even some of the catalysts involved in electron transport, have been disheartening. DNA origami could provide the scaffold to hold proteins in place, says Hao Yan, a biochemist at Arizona State University in Tempe. As part of a collaboration funded by the US Department of Energy at the university's Center for Bio-Inspired Solar Fuel Production, he has been looking to use DNA origami as the basis for an artificial leaf that makes hydrogen fuel from water.

Yan plans to use a cage-like DNA structure to position the proteins and to bind manganese, a crucial component of the water-oxidation process. The goal is to better direct electron flow in an artificial system that requires precise placement of components. "If we can really control all the electron-transfer sites then we can improve the efficiency," says Yan.

Put a drug in a box

Last year, Kurt Gothelf, director of the Centre for DNA Nanotechnology at Aarhus University in Denmark, and his colleagues, reported that they had made a box from DNA origami3. One strand of DNA holds the lid shut; a separate DNA 'key' springs it open. The invention prompted many to wonder what could be put into the box and subsequently released.

William Shih, a DNA nanotechnologist at Harvard Medical School in Boston, Massachusetts, is keen to exploit this kind of vessel for drug delivery, but the challenges are significant. Getting the box to pass through a cell membrane is going to be difficult, he says. He proposes covering the box with a membrane similar to those sported by some enveloped viruses.

These viruses also have special proteins to facilitate entry. Shih says that he can take viral proteins or related proteins and fix them to the outside of his DNA cages, but he faces a long list of challenges. "There's a lot of stuff," Shih says. "At this point it's just a concept."

Go for gold

Many have talked about using DNA origami as a substrate for nanoelectronic circuitry, such as in plasmonic devices.

Plasmonic devices couple light waves with charges on a metal surface and offer the speed of information transfer that light provides, but at sizes smaller than those to which technologies such as fibre optics are limited. Current lithographic techniques run into trouble when trying to arrange metallic materials such as gold into patterns with features smaller than about 100 nm in size. Rothemund says that gold spheres might be positioned using DNA origami to make structures with better optical qualities.

Now we've got DNA origami lined up like little ducks in a row. ,

He has already taken steps, in concert with IBM Almaden in San Jose, California, towards arranging metal nanoparticles. Last year they managed to arrange DNA triangles on a lithographically patterned surface4. Jennifer Cha from IBM and her colleagues subsequently showed that they could place gold nanoparticles smaller than 10 nm onto each origami structure by affixing strands of DNA to the gold that were complementary to loose ends on the DNA triangles5. The work is still quite crude, says Rothemund. They haven't got an active device yet. "Much of the next five years will be spent perfecting techniques to place DNA origami on surfaces where we want them and making this technique widely available to other scientists," he says. Importantly, they've tacked down the shapes rather than having them "bobbing around like jellyfish in solution", says Rothemund. "I didn't think anyone would solve that problem in ten years. Now we've got DNA origami lined up like little ducks in a row."

Pushing past smileys

The grand schemes go on. Researchers imagine an artificial ribosome capable of building custom enzymes, a matrix for supporting artificial organs, or a DNA origami network designed to support a neuronal network connected to electrical circuitry. Researchers are attempting all of these, but they will face some serious hurdles. Buffer solutions must be finely tuned, otherwise structures fall apart. And large, complicated structures can take up to a week to fold completely.

Change is needed to the basic, almost slapdash synthesis approach of using unpurified DNA that isn't adequately sequenced, says Shih, "just because we got away with it so far doesn't mean we want to get away with it forever".

But as long as this growing group of researchers keeps trying to make different shapes, other applications will appear. Rothemund can't predict quite how, though. "The problem is that you can do all these little cool things but they in no way form a whole system. It's not exactly clear what kinds of systems we're going to be able to build."

He's working hard to find out. "In the past six months, I've mostly been back at home, coding in my underwear again, which hopefully means good things are happening."

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