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首页> 外文期刊>ACS applied materials & interfaces >Fabrication of Inverted High-Density DNA Microarrays in a Hydrogel
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Fabrication of Inverted High-Density DNA Microarrays in a Hydrogel

机译：水凝胶中倒进的高密度DNA微阵列的制备

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摘要

Current techniques for making high-resolution, photolithographic DNA microarrays suffer from the limitation that the 3' end of each sequence is anchored to a hard substrate and hence is unavailable for many potential enzymatic reactions. Here, we demonstrate a technique that inverts the entire microarray into a hydrogel. This method preserves the spatial fidelity of the original pattern while simultaneously removing incorrectly synthesized oligomers that are inherent to all other microarray fabrication strategies. First, a standard 5'-up microarray on a donor wafer is synthesized, in which each oligo is anchored with a cleavable linker at the 3' end and an Acrydite phosphoramidite at the 5' end. Following the synthesis of the array, an acrylamide monomer solution is applied to the donor wafer, and an acrylamide-silanized acceptor wafer is placed on top. As the polyacrylamide hydrogel forms between the two wafers, it covalently incorporates the acrydite-terminated sequences into the matrix. Finally, the oligos are released from the donor wafer upon immersing in an ammonia solution that cleaves the 3'-linkers, thus freeing the oligos at the 3' end. The array is now presented 3'-up on the surface of the gel-coated acceptor wafer. Various types of on-gel enzymatic reactions demonstrate a versatile and robust platform that can easily be constructed with far more molecular complexity than traditional photolithographic arrays by endowing the system with multiple enzymatic substrates. We produce a new generation of microarrays where highly ordered, purified oligos are inverted 3'-up, in a biocompatible soft hydrogel, and functional with respect to a wide variety of programable enzymatic reactions.
机译：用于制备高分辨率的电流技术，光刻DNA微阵列患有限制，每种序列的3'末端锚固到硬质基质上，因此对于许多潜在的酶促反应是不可用的。在这里，我们展示了一种将整个微阵列转化为水凝胶的技术。该方法保留了原始图案的空间保真度，同时除去所有其他微阵列制造策略所固有的错误合成的低聚物。首先，合成供体晶片上的标准5'-up微阵列，其中每个寡聚聚焦在3'末端的可切割接头和5'末端的氨基磷酰胺锚定。在合成阵列之后，将丙烯酰胺单体溶液施加到供体晶片上，并将丙烯酰胺 - 硅烷化受体晶片放置在顶部。作为两种晶片之间的聚丙烯酰胺水凝胶在两种晶片之间形成，它共价掺入基质中的丙烯酸封端的序列。最后，在浸入氨溶液中，寡核苷酸释放在氨溶液中，从而使3'-接头的氨溶液中，从而在3'末端释放寡核苷酸。阵列现在在凝胶涂覆的受体晶片的表面上呈现3'。各种类型的凝胶酶反应证明了通过赋予多种酶底物的系统而言，可以容易地构造比传统的光刻阵列更容易构造得比传统的光刻阵列更远。我们生产新一代微阵列，其中高度有序，纯化的寡核苷酸在生物相容的软水凝胶中倒置3'-up，并且相对于各种可编程酶促反应的功能。

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