my name is William shei I'm an associate professor of biological chemistry and molecular pharmacology at Harvard Medical School Dana Farber Cancer Institute and the VIS Institute for biologically inspired engineering it's my pleasure to share with you today some recent technical advances in the field of structural DNA nanotechnology from my laboratory and those of my colleagues we're all familiar with the biological role of DNA as a information repository principally for coding for protein sequence and for regulation of protein expression and I'm not going to speak about that at all today instead I'll be talking about using DNA itself as a building material and harnessing that in order for us to construct nanoscale objects for example shown here is an electron micrograph of actin filaments that are about 7 nanometers in diameter and Below we can see this peculiar Pac-Man shaped object this is a a structure that was built entirely from DNA and we designed it for the purpose of taking bite-sized chunks out of the actin filament while this project hasn't yet been successful however hopefully this image drives home the complimentarity between the dimensions of our design DNA Nano structures and biological macromolecular complexes so I've been working in this field for over over a decade now and I'm continually surprised by advances in the field I have these preconceived notions of the limitations of DNA and they're always Shattered by the latest new discovery and today I'm going to be sharing with you in the first two sections respectively two recent developments that enable us to design and assemble DNA Nano structures of the size and complexity of this object shown here about 30 nanometers in diameter one of the most important goals for DNA nanotechnology is to self Assemble ever increasing objects of ever increasing complexity over time for example is it possible that within the next decade or two that we can self assemble objects that are let's say a thousand times as complicated that have a thousand times as many unique components as the object shown here a second question is what are these things good for and in the third part of this lecture I'll be discussing some applications in my laboratory uh as tools for molecular biophysics and tools for future Therapeutics where we think these objects might prove useful we're inspired from natural systems we know that they can carry out many amazing behaviors they can build they can adapt they can heal uh they can reproduce and these are capabilities that human technology struggles to reproduce on on any kind of length scale but what's especially remarkable is the ability of life to carry these out on the molecular scale so for example here on the left is a picture of a ribosome and this is of course a machine that's about 25 nanometers in diameter that takes information encoded in messenger rnas and then translates that into a specific sequence of amino acids to produce a polypeptide quite amazing machine on the right we have the T4 bacteria Fage it's a bit larger in length scale we can see that the viral capsid itself head capsid is about 100 nmet in diameters it looks like a little nanoscale hypodermic syringe that docks onto the surface of its spectal host and then injects its DNA cargo into the cell against an osmotic pressure gradient of course what makes this all possible is that uh living systems have invented molecular manufacturing and they've come up with a very robust and clever way of doing this they of course synthesize these biopolymers they could be polypeptide chains polynucleotide chains that then self assemble into the desired structure now if uh somebody asked you about 10 years AG ago would it be possible to generate an object of this complexity using some kind of human-based Technology most people would have been skeptical and yet I'm going to show you a new technology DNA nanotechnology developed in the last especially with advances from the last 10 years that now make it possible for us to self assemble in a programmable way structures of the same kind of complexity as you see here not yet of the functional complexity but nevertheless we we think this is a encouraging first step because uh along the pathway to this kind of functionality we first need to master structural complexity we're going to be using DNA as our building material and we know that DNA very similar to proteins and other macromolecules from Life are very complicated molecules there's many different atoms but it turns out the key point for DNA nanotechnology is that the robust base pairing properties of DNA allow us to AB ex ract away those chemical details which is going to make the act of Designing the Nan structures much simpler and in fact there's only three characteristics of the DNA that we need to remember for the purpose of DNA nanoc construction one is that it's a ladder with antiparallel strands secondly there's a right-handed twist for bdna and we need to know that the twist is around 10 and a half base pairs per turn turns out we can switch that around a little bit and finally uh we need to know that a pairs with t c pairs with G and anytime you deviate from that pairing you're going to destabilize the structure and it's because the propensity of DNA to form this very regular structure enforced very strictly according to this Watson CC Bas pairing that gives us its power in being able to generate these large structures with very little design work the father of the field of DNA nanotechnology is Ned Seaman and NYU he invented this field about 30 years ago his training is as a crystallographer and the way he came up with the idea is as follows he was sitting in the campus Pub one day just drinking his beer and suddenly what popped into his head was this wood cut from MC eer he had been collaborating with some of his friends on uh DNA holiday Junctions and he had his Eureka moment why not replace the flying fish with DNA holiday Junctions the notion was that if he could rationally design a porous Crystal out of DNA and then he could take the target protein he was interested and then dock that into each unit cell into a stere stereotyped orientation then he'd able be able to impose that Crystal in order on the target protein and therefore make the X x-ray crystalogy problem easier for these large macro molecules that are otherwise difficult to crystallize uh he's been working on this problem for over 30 years it's an important goal and he's made some interesting progress I'll have more to say about that in the third segment but in the meantime he's had some interesting Landmark successes the first really noteworthy advance that he reported came out in 1992 or so in nature so this is a DNA Cube where each edge of the wireframe cube is two turns of a double helix each face is a circular strand of DNA and the entire object has dimensions of about 10 nanometers uh so the first time I think people saw this they thought well this is really cool but that doesn't look like biology to me and what I hope to convince you today is that this is in fact an extremely powerful techn technology yes it's fun but it's actually potentially very useful as well for many different applications of course if we're building from DNA strands and we're just making double heles then that's boring the power of DNA nanotechnology is that we can build with Branch Junctions with the previous example the cube each one of those vertices is a three branch Junction but it turns out the most powerful Motif so far in structural DNA nanotechnology has been a for branch Junction a holiday Junction so on the upper left here we have a schematic using simple letter notation of the Strand so we can see the Canan strand starts five Prime CC G goes to its three prime end and if you look closely at this you can see that there's four different sequences and they have the proper sequence complement par it in order to generate a holiday Junction that actually is immobile it can't Branch migrate due to its sequence and we know from structural studies that this object likes to stack into two double heles that are connected at a joint and it turns out this is really the building block that's been the most fruitful for DNA nanotechnology so the idea is as follows if you only have one holiday Junction now you have two Hiles that can wobble around in order to fix those two heles to make a rigid building block what we do is we simply introduce a second holiday Junction Downstream and now when we fix that those two double Hiles with two holiday Junctions we have that rigid building block that we want now with four sticky ends the next step is to build two versions of this building block in this example we have a red one and we have a blue one and we design the sticky ends with the following complimentarity in this example so let's say we make the sticky end on the upper right hand side of the red block and we make that compatible with the lower left hand side of the Blue Block and so on and so forth in order to create this kind of checkerboard fashion hopefully you can see that we would be able to self assemble these two bricks into an infinite two-dimensional lattice as shown below I don't have the experimental images for this uh but it suffice to say that this method actually worked it's quite amazing you can design a two-dimensional Crystal the step after this would be to say well instead of just two bricks two tiles what if I had 10 tiles or what if I had 100 tiles can I now make non-periodic structures that are highly complex just with self assembling tiles and unfortunately nobody has really demonstrated this method uh extending this particular method to hundreds of tiles although what I'll show you shortly is is one method DNA origami that can achieve this kind of complexity and in the second segment uh something called single stranded bricks that can do something very similar to what I just uh prescribed the method of DNA origami is a particular flavor of structural DNA nanotechnology it was developed by Paul Rodman at Caltech he published this in 2006 and the basic idea is as follows so imagine you have a long single strand of DNA the 7,000 base Genome of the M13 bacteria phage that's the gray strand in this animation we know what the sequence is and based on that known sequence we chemically synthesize hundreds of short olle tides that are 20 to 60 bases long that are programmed by Watson cric complimentarity to pinch that long strand into a parallel array of helices after heating everything up to about 65° centigrade and then cooling down to room temperature over the course of an hour at the end of the assembly you end up with this parallel array of double heles where adjacent double Hiles are held together by these holiday Junction crossovers that I described to you a couple slides ago so this is a half crossover and then here we have a full DNA crossover importantly what you just saw was an animation not a simulation in fact we have a very poor understanding of the order of events of folding these objects we just know that if we program them in a way where all of the scaffold ends up base paired to staple strands then we have an extremely high probability of forming the desired structure so it's a very active area of research for us to try to understand better the mechanism of folding and we we hope hope that will actually help us to to design more complex structures in the future so Paul Rodman used this method in 2006 to make structur such as this disc with three holes it has dimensions of about 100 nanometers by 100 nanometers by 2 nanometers this is atomic Force micrograph the example in the upper left hand corner represents in size just part of the upper lip of the object so this is quite large by macro molecular standards it's like we have two ribosomes worth of molecular Silly Putty that we can Mash into any desired two-dimensional cookie cutter shape one of the very interesting things that he pioneered was that he developed a way to make this din origami where he made each one of those staple strands in two different flavors so one flavor just made the structure as you saw the second flavor had the identical sequence but had a surface feature a dumbbell that's sticking out of one of its ends and so what that means is anytime he used the original flavor and he added it to the folding mix then you'd get a plain vanilla din origami surface at that at that location but then if you replace that sequence with the longer sequence the one with the feature now you get that same shape but a bump over that feature and in that way he can see that this rectangular din origami could be treated as a molecular braid board where let's say it has 200 different positions we can decide at each position whether or not we want to create a bump or have no bump in effect we have something that's like a bit map and we can create new patterns simply by repiping different patterns of the no bump and plus bump strands for each one of the locations so for example here we can see that he's designing something that will say DNA and have a little picture of DNA these structures actually become very sticky at their ends because they have lots of blunt ends and then they'll make a continuous ribbon that says DNA you can see that he made a map of the Americas he's a very humble guy so he apologized to the rest of the world for stopping at the Americas but um DNA is a little bit expensive so he he stopped at the moment maybe by now he's made the rest of the world uh and you can you can program them to link up in specific ways and in that way you can self assemble two-dimensional crystallin objects so what about getting to three dimensions as I alluded well we can get our initial inspiration from macroscale paper origami where we know we have we're quite familiar that if we fold flat paper in many ways we can get quite intricate three-dimensional shapes so this is the famous Crane and if you're really diabolical like Robert Lang then you you might note that uh if you can fold these papers in very in especially intricate ways then you can make incredibly complicated objects that we we can see some examples of here now nothing I'm going to show you with DNA is as complicated as this but again as I mentioned one of our goals is to scale to ever increasing complexity so we hope that someday we actually can self assemble DNA into objects of this kind of complexity so that group in Denmark that I just mentioned e Anderson yur Kims K goal they were able to design that M13 to fold into six different sheets and then they program those six sheets to fold up into a threedimensional box with a hollow inside they designed a lid that can open in response to some kind of molecular key so this would is the first example of three-dimensional Hollow D origami so where my group wanted to contribute was to make solid three-dimensional D origami structures and the idea is as follows so first of all we we know that we can curl up DNA due to the helicity of the DNA heles and I'm going to go through a little thought experiment just to give you a flavor of what this is about so here we have on the far left three double heles that are ranged into a little the origami you can see if you look closely they're connected by those holiday Junction crossovers to keep the hes parallel and in this Arrangement it's making a flat sheet of three Hiles so now imagine what would happen if we moved these crossovers on the top two base pairs to the left then that's going to move that double helix behind the plane of the page and likewise if we move those two crossovers two base pairs to the right that's going to move that double helix in front front of the plane of the page the take-home message here is that simply by shifting around the position of those crossovers with respect to each other we can achieve curvature of these DNA origami sheets along the axis of the double heles so that's the first key so now let's extend that and build a actual solid threedimensional structure so here we have another representation of a d origami where each one of these cylinders represents one of those double Hiles so it's similar to the example in the upper left but now just rotated into the dis orientation so this would represent the pattern of the scaffold running through those heles but for the purpose of this explanation I'm going to leave that invisible it's there I'm just not going to talk about it that or the staple strands and so what we're going to do is we're going to shift around the position of those crossovers so now these heles no longer pref prefer to be planer but instead prefer to curl up into some kind of specific geometry and in this example what we're doing is we're trying to curl up the structure into a corrugated s shape furthermore anywhere we where we have the orange sheet that touches the white sheet touches the blue sheet we're routing those staple strands through those interfaces so for example we might have a staple strand that starts seven base pairs on this Helix then goes seven base pairs here seven base pairs seven base pairs seven seven base pairs and in that way if the structure forms the way we intend it to it should be highly cross L by the staple strands that are traversing the different heles so looks good on paper okay what happens in the test tube when we tried it and perhaps who can say of course when we threw all the strands together and tried to fold the object then it didn't work we got a pile of molecular spaghetti that we could see under the electron micrograph um but we didn't want to give up and eventually H Deets in the group came up with a key Insight which is it's not that these three-dimensional objects now are unstable they're dynamically simply they're more difficult to achieve kinetically and so what we found is that we could only get appreciable yields of these objects if we folded them instead of for an hour going from 65 room temperature if we folded them for more like a week then we could start to get appreciable yields of the objects ranging from 10 to 50% yield so we can see here uh one of the objects that was built by Shan Douglas instead of three layers it was with 10 layers and then we have the electron micrographs below we can see that we get a close resemblance between what we observe in the electron micrograph and the or projection orientations of our design structure this is work that we published back in 2009 in the meantime our group and others have been hard at work trying to improve the method so the important thing here was that we we could get something to fold it all and now we're trying to get better yields improve the folding times so there have been a couple of important discoveries since then one has come from hendrik de's Lab in Munich where they've discovered that these structures tend to have a favored temperature which they fold faster than the other temperatures so instead of spending the same amount of time at 65 degrees down to room temperature for example the structure maybe folds faster at 50° Centigrade so what they found is if they do most of their folding at 50° Centigrade then they can get it to fold maybe in order of magnitude faster which uh makes a a lot of improvement for our Our Lives as scientists designing them they also suffer less thermal damage with a slower folding ramp we've also learned some details about how to design the the strands the Crossovers the break points that I don't have time to go into in this presentation but I encourage you to look at some of our Publications if you want to see the latest discoveries and how to make this process work better so now I'm going to go through a panel a gallery of different objects built using this method by our laboratory to give you a flavor of the generality of the method so the example on the top is what I just showed you we call it the monolith it was built by Shan Douglas you might say that it looks a little bit like a nanoscale crystal honeycom crystal but it's important to keep in mind that every element of the object is associated with a unique sequence and therefore is independently addressable this is quite different from most nanop particles that we see in in synthetic nanotechnology today the example on the bottom was built by Francisco gra we call it the Genie bottle we called it that because uh in one version not shown here we only folded part of the M13 scaffold and the rest of it was coming out of the lip of the object kind of like wisps of smoke these are all 20 NM scale bars so here again 20 nmet scalebars on the left is the object built by Shan Douglas we call it the square nut it has a 7 nanometer hole in the middle it has a front end and a back end and if we make the sticky ends on the front end compatible with the sticky ends on the back end then we can self assemble filaments that are somewhat reminiscent of actin filaments or microtubules although in this case they don't yet demonstrate any Dynamics they're just equilibrium formation of these long polymers on the right is the object built by Tim leele we call it the rail Bridge again every cylinder is one double helix and we can see as we go through cross-sections of the object we have a different arrangement of double Hiles and we can understand from this example that it is kind of analogous to sculpture that you could imagine the sculpture begins with a solid block of marble in our case these parallel rays of double heles and in design space we're chipping away at that solid block until we achieve whatever threedimensional structure we actually want in relief once we have our final design then we uh what we're doing is we're compiling that threedimensional structure into a series of DNA strands that are going to self assemble with the M13 scaffold into that object here's the object built by Jan hogberg when he was in the group we call the slotted cross I'll have more to say about this object in the next slide this is another crossed object we call the Stacked cross built by hend Deets again these are all 20 NM scalebars this one looks a little bit like stacked molecular celery we even designed a little molecular cavity on the top where where we initially imagined we could host protein guests on the inside of that cavity so let's take a closer look at that slotted cross from bjor herberg so here what he's done is he's generated animation where he stylized the routing of the scaffold strand through the structure it's designed as a h domain and an O domain and the middle of the H domain is designed to pass through the middle of the O domain and it's all folding from just one long en 13 scaffold I was quite amazed that this thing folded at all um but the yields are not so great at the moment just a few percent so now what I'm doing is I'm zooming in on what we call the Strand diagram that describes a blueprint of the object it's like we take all the helices and then we spay them out onto a two-dimensional surface and in this case the blue represents that M13 scaffold Strand and those colored strands represent the staple strands and this part of the object is the upper left hand corner of the H domain and so we can if you look closely you can see that the staple strands what they're doing is they're binding to part of the scaffold Strand and then they're crossing over to a different part of the scaffold strand to pull those uh components together to make the three-dimensional shape we can zoom out and then here you can get an appreciation that it is kind of like a blueprint you can make out which part is the H domain which part is the O domain and if you look closely you can actually actually see where the H domain and O domain are being connected by that long scaffold strand all of the examples that I've shown you so far have been used built using this honeycomb lattice Paradigm where we're using these corrugated sheets it turns out that more naturally fits the preferred Twist of DNA of 10.5 base pairs per turn but it turns out we can also self assemble these DNA sheets in a square lce format uh the only Proviso is that now we have to underwinding and quite interestingly what happens is the structure will still form but it then compensates by having a global super twist in the in the right-handed Direction so it's quite analogous to how plasma DNA for example will have a right-handed super twist when when it's underwound as as we find uh in most cells one very important development in the field is uh software with a graphical user interface to make it accessible to people who are outside the field but also just to make the process faster more robust and convenient for experienced practitioners so uh for this really powerful software suite called CAD Nano we or thanks to Sean Douglas he developed this software when he was was a graduate student in my group now he's a assistant professor at UCSF at the time of this filming so I encourage you to check out the software he's continually improving it at CAD nan. org and what we can do is now again with a graphical user interface within an hour or so we can design different shapes and then compile that into the sequence of DNA strands that can self assemble into that object what if you wanted to build structures well the most obvious idea is to just get more parts so you can remember as as a kid the first time you got a Lego Lego set it was enthralling but then about two hours later you now were hungry for additional Lego pieces so that's the big drive for our field can we get more Lego pieces into the structure but in the meantime we can do other things that will allow us to get a little bit bigger so one example is just a build with wireframes that have high strength to weight so in this example what we've done is we've added staple strands that fold that M13 scaffold into this wireframe structure each one of these struts in this example is a six helix DNA N Tube and then we designed sticky ends such that they're compatible and we can get this structure this zape structure to fold into a double triangle with now 10 of these six celix bundle termin each with a unique set of sticky ends in this example what we did is we programmed three separate double triangles to form in three separate test tubes and we programmed it to form this network on the bottom this is a Schlegel diagram and for those of you who might recognize this you might see that this is actually a schle diagram for a wireframe icosahedron this object has a overall diameter of about 100 nanometers each one of the struts has a length of about 45 naners and here on the lower left hand corner we can see an animation macroscale animation reenactment of the self assembly of these double angles into a wireframe icosahedron what we find is that this process works in the test tube as well no hands required so again what we do is we fold each of the double triangles in three separate test tubes we then mix them together to form the desired wireframe object so let's take a look using electron microscopy so here we see with the one micron scale bar we see a bunch of of objects that seem to have about the right size about 100 nimet in diameter there's Aggregates as well so the self assembly is not perfect but we're glass half full kind of folk we're encouraged by something that works uh even partially so now we've zoomed in you have a 500 nmet scale bar and we can tell that there's some kind of wireframe action going on zoom in some more now we have a 200 nmet scale bar and it's it's starting to look like the wireframe structure that uh we imagined of course you have some misassemblies as well and then now if we go to the highest effective magnification for this negative stain method 100 nanm scale bar we can see that objects in fact they look like they have lots of these triangular faces they look like they have five-fold vertices and we're able to make an object that now is something like five times the mass of a ribosome it has overall dimensions the size of a of a mediumsized virus and this is all just powered by Watson C base pairing a pairs with t c pairs with G uh it's remarkable that we can push it this far but we're greedy and we dream about being able to extend this to objects that are a thousand times more complex or even more than that someday another kind of wireframe structure from macroscale engineering that inspired us are these floating compression sculptures from the artist Ken selson and the idea here for these sculptures is that you have these beam that are bearing compression that aren't touching each other directly but instead they're connected by cables that are bearing tension and if you wire this up in in the correct way it's then it's a balance between the tension of the cables and the compression of the beams and you end up with the object that has high strength to weight and has other interesting features for example if those cables have some elasticity then if you put a global force in the object then uh it will deform and every individual uh strut will rearrange in threedimensional space when you that will relieve that stress then it'll bounce back to the original shape so we wanted to see if we could implement this using DNA origami this is work that was led by Tim leel and Bon hogberg when they're in the group in collaboration with Don inber so what they did was to design those staple strands to fold this M13 scaffold into three different uh struts each of these struts in this cases 13 Hiles it's actually grabbing three separate segments of that scaffold in order to make each one of those uh 13 Helix struts so we again add everything together heat it up cool it down and remarkably enough you can form structures like this in the test tube in fact we started to play games about looking at how much stress we could put the objects under and have them still fold so what happens is that you have these single stranded DNA elements that are acting like entropic coils they're exerting tension and if we simply design those uh cables to be shorter have fewer number of bases then it's going to exert a larger force over the same design distance between the two compressed elements and what we find by found by continually shortening these cables is that we could self assemble the structures up to about 14 PE Newtons of force that was the the calculated Force for the shortest cable that we're able to self assemble the objects in other words we're able to self assemble these DNA objects against twice the force that's can be generated by powerful set skeletal Motors such as kinesin or mein this is all powered by just DNA based pairing we we believe that these kinds of structures may prove useful for applications in tissue engineering and regenerative medicine so of course cell biologists have noted for a while now that cells uh especially going through development can communicate with their outside environment with each other using mechanics so they might pull on The extracellular Matrix and The extracellular Matrix may pull back uh and you might have by introducing deorations into the extracellular Matrix or within the within the cytoskeleton of the cell you can then trigger biochemical events so we envision a day where we can use these kind of DNA n structures that can deform in response to some kind of mechanical stress and then translate into a biochemical event it could be release of a growth factor or maybe it could involve catalysis of some kind of chemical reaction so we believe that this could be useful for regenerative medicine so the last thing that I'd like to show you for this section is work from hendl Deets Sean Douglas assisted on this work everything that I've shown you thus far has involved double huses that are straight and Hendrick wanted to ask the question could you build structures curv structures where the Hiles now are falling an arc instead of going straight and the basic strategy for implementing this is as follows so here we have again every cylinder represents a double helix these planes that are slicing through the double helixes represent the positions at which those crossovers are occurring so it turns out in this example they're only occurring every seven base pairs and he asked the question well what would happen if he replaced the double helical segments on the top so the the orange segments with shorter double helices that only have six base pairs between planes and what if we replace the Hiles on the bottom the blue ones instead of seven base pair segments he had eight base pair segments so mechanically now on the top those elements are going to be under under tension because you have less material in the same amount of space they're going to be stretched out the heles on the bottom are going to be under compression because we now just stuffed more material into the same amount of space and the system is under stress and so it's going to relax of course by bending so this is the way to relieve that tension on the top and compression on the bottom does this actually work when we attempt this in the test tube and the answer is yes so hendrik implemented this using an 18 helix DNA bundle that's Illustrated on the left hand upper left hand side and so what he did was he had a stereotyped straight region these white regions and then he had a experimental region that's indicated here in Red so that's where he's going to be introducing those longer and shorter elements to induce the bending of the structure we can see for the control you get this nice rigid straight object so what happens when he introduces some small number of shorter strands double heles on the top and then longer ones on the bottom he could get a reliably predicted 30° Arc at that position if he has roughly twice the number of perturbations then you can get to 60° angle kept on going you get 90° you can get 120° angle that's quite remarkable this is now getting down to a 10 NM radius of curvature but then he kept on going and he found he could go all the way to 180 Dees in this example so this is something that has a 6 nmet radius of curvature it's comparable to the tightness of wrapping of DNA double helices around histones in a nucleosome so in that case that's powered by protein DNA interactions in our case this is powered by DNA Bas pairing interaction CS so here what we have is an animation prepared by Shan that explains the bending principle so again what we're doing is we're introducing more double more base pairs or longer double heles on the left and then shorter ones on the right and we can see a little graph on the lower left hand side that tells us how many base pairs per turn that we have for each of these different elements and at the most extreme example we're actually getting 15 base pairs per turn on the left which is severely underwound and only six base pairs per turn on the right which is severely underwound and I was quite flabbergasted that it should be possible for us to torture DNA to this extent now in fact once you get to those extremes our folding yields do start to go down so we can see that we're we're at the edge of what we can do to DNA but still it's quite remarkable that DNA is is so robust that the 10.5 base pairs per turn is simply what it PR first to do but if you put enough stress on it you can make it do things that deviate from that ideal by quite a bit so HCK and Sean now use the method to make a variety of different structures so on the upper left hand corner we have a six hexd bundle that's folded into a series of uh half of 180° arcs of increasing radi curvature so you make a spiral on the lower left hand corner we have an object that's programmed to self assemble into a beach ball out of six heix pundles we can see objects that are making concave triangle this is designed by Shan Douglas and then here we have those 120 degree arcs that are repurposed so we made sticky ends on the two ends of this little Boomerang to be complimentary so that you can have three identical versions of them will come together to make a larger triangle so in conclusion hopefully I've persuaded you that DNA origami is a highly versatile method for building both two-dimensional and three-dimensional structures of quite remarkable complexity about twice the mass of a ribosome where we're moving to next is to try to build structures that are more complicated you might wonder what's preventing us from building something a thousand times larger already today and the main problem is that we have errors in the self assembly and for example for one of these objects we might have a yield in best cases 75% or so which might sound pretty good but now if you wanted to build an object that's a thousand times bigger then you might argue that the probability if you just mix these things together a thousand of them together the probability that all one that none of the 10,000 would have any defect would be 75 to the 1,000's power which if you do the math that's basically zero so there's a lot of activity in the field trying to improve the Fidelity of this self assembly uh other methods like hierarchical self assembly error correction that'll allow us to scale Mount complexity and build really uh really very complicated objects of the future