复杂的二维形状的从单链DNA的瓷砖自组装
Summary
DNA tiling is an effective approach to make programmable nanostructures. We describe the protocols to construct complex two-dimensional shapes by the self-assembly of single-stranded DNA tiles.
Abstract
Current methods in DNA nano-architecture have successfully engineered a variety of 2D and 3D structures using principles of self-assembly. In this article, we describe detailed protocols on how to fabricate sophisticated 2D shapes through the self-assembly of uniquely addressable single-stranded DNA tiles which act as molecular pixels on a molecular canvas. Each single-stranded tile (SST) is a 42-nucleotide DNA strand composed of four concatenated modular domains which bind to four neighbors during self-assembly. The molecular canvas is a rectangle structure self-assembled from SSTs. A prescribed complex 2D shape is formed by selecting the constituent molecular pixels (SSTs) from a 310-pixel molecular canvas and then subjecting the corresponding strands to one-pot annealing. Due to the modular nature of the SST approach we demonstrate the scalability, versatility and robustness of this method. Compared with alternative methods, the SST method enables a wider selection of information polymers and sequences through the use of de novo designed and synthesized short DNA strands.
Introduction
以前核酸自组装工作1-25已导致各种复杂的结构，包括DNA 2的成功构建– 5,8,10 – 13,17,23或RNA 7,22周期性3,4,7， 22和算法5二维晶格，彩带和10,12管4,12,13，3D晶体17，11多面体和有限，2D图形7,8。一种特别有效的方法是脚手架的DNA折纸，据此，单一支架链是由许多短的辅助订书钉链折叠以形成复杂形状9,14 – 16,18 – 21,25。
我们最近报道的方法构建与使用单链砖（SST）规定的2D形状离散纳米结构，并表现出与结构的复杂性堪比DNA折纸26。这article是我们以前的工作26的改编，并介绍安排单独寻址到自助服务有限精良2D形状与精确规定的尺寸（宽度和长度）和形态的详细方案。该SST方法的一个关键优势是其模块化。结构的每一个组成部分SST作为在装配模块化施工单位，而这些表层海水温度的不同子集产生不同的形状。因此，我们建立构造纳米结构与规定的尺寸和形状，从短，合成DNA链一个通用的平台。
自助服务包含四个畴，每次10或11个核苷酸长（ 图1A）。该表层海水温度结合使得它们平行螺旋通过创建交叉联系在一起的DNA点阵。各交叉是结构域2和3的磷酸盐在图中为清晰起见人工拉伸之间的磷酸盐。该交叉间隔两个螺旋形圈（21个碱基）分开（<STRONG>图1B）。该复合矩形在螺旋和螺旋的圈数提到了它们的尺寸。例如，一个矩形为六螺旋宽和八个螺旋变为长被引用作为6H×8T矩形。海表温度可以被排除在外，添加或以其他方式重新安排，以创建任意形状和大小（ 图1C）的结构。例如，矩形的设计可以卷成管具有所需长度和半径（ 图1D）。
可替代地，该矩形的SST晶格可以被看作是一个分子帆布组成的SST的像素，通过为7nm每3纳米。在这项研究中，我们使用的310全长内自助服务一个分子帆布，24全长自助服务构成的左和右边界，及28半身自助服务形成的顶部和底部的边界。画布具有由交叉链接24双螺旋和每个螺旋包含28个螺旋转角（294碱基），并因此被称为一24H×28T直角的画布。所述24H×28T帆布具有类似于从M13噬菌体脚手架创建的DNA折纸结构的分子量。
Protocol
Representative Results
Discussion
在结构形成步骤中，以保持镁阳离子的适当的浓度是很重要的（ 例如 ，15毫摩尔）的DNA链混合物自组装的DNA纳米结构。同样，在琼脂糖凝胶表征/纯化步骤，以保持适当的镁阳离子浓度是很重要的（ 例如 ，10毫摩尔）在凝胶和凝胶运行缓冲液电泳过程中保持该DNA的纳米结构。为24H×28T矩形结构，我们在不同的Mg ++的浓度下测试退火，发现通常形成在10的范围内的结构- 30mM的Mg <s…
Disclosures
The authors have nothing to disclose.
Acknowledgements
这项工作是由海军研究青年学者计划奖N000141110914，海军研究基金N000141010827局，美国国家科学基金会职业奖CCF1054898，美国国立卫生研究院主任的新的创新奖1DP2OD007292办公室和威斯研究所生物启发工程学院启动基金（以PY）出资清华 – 北京中心生命科学基金启动（到BW）。
Materials
| Name of Material/ Equipment | Company | Catalog Number | Comments/Description |
| DNA Strands | Integrated DNA Technology | Section 3.1 | |
| SYBR Safe DNA gel stain | Invitrogen | S33102 | Section 3.4.2 |
| Freeze'N Squeeze DNA Gel Extraction Spin Columns | BIO-RAD | 731-6166 | Section 3.6 |
| Bruker's Sharp Nitride Lever Probes | Bruker AFM Probes | SNL10 | Section 4.3 |
| Safe Imager 2.0 Blue Light Transilluminator | Invitrogen | G6600 | Section 3.6 |
| Centrifuge 5430R | Eppendorf | 5428 000.414 | Section 3.6 |
| Transmission Electron Microscope | Jeol | Jem 1400 | Section 7.4 |
| Multimode 8 | Veeco | Section 4 | |
| Typhoon FLA 9000 Laser Scanner | GE Heathcare Life Sciences | 28-9558-08 | Section 3.5 |
| Ultrapure Distilled water | Invitrogen | 10977-023 | Section 3.7.1 |
| Mica disk | SPI Supplies | 12001-26-2 | Section 4.1 |
| Steel mounting disk | Ted Pella, Inc. | 16218 | Section 4.1 |
| carbon coated copper grid for TEM | Electron Microscopy Sciences | FCF400-Cu | Section 7.2 |
| tweezers | Dumont | 0203-N5AC-PO | Section 7.31 |
| glow discharge system | Quorum Technologies | K100X | Section 7.2 |
| DNA Engine Tetrad 2 Peltier Thermal Cycler | BIO-RAD | PTC–0240G | Section 3.3 |
| Owl Easycast B2 Mini Gel Electrophoresis Systems | ThermoScientific | B2 | Section 3.4.3 |
| Seekam LE Agarose 500G | Lonza | 50004 | Section 3.4.1 |
| GeneRuler 1kb Plus DNA Ladder, Ready-To-Use 75-20000bp | ThermoScientific | SM1333 | Section 3.4.4 |
| Nanodrop 2000c UV-vis Spectrophotometer | ThermoScientific | Section 3.7 | |
| 0.2 um filter | Corning Inc. | 431219 | Section 7.1.2 |
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