无需逆转录的折叠RNA文库的DNA展示实现RNA-SELEX
DNA display of folded RNA libraries enabling RNA-SELEX without reverse transcription.
作者信息
MacPherson I S, Temme J S, Krauss I J
机构信息
Department of Tropical Medicine, Medical Microbiology and Pharmacology, John A. Burns School of Medicine, University of Hawai'i at Manoa, 651 Ilalo St., Biosciences Building, Suite 325, Honolulu, Hawaii 96813-5525, USA.
Department of Chemistry, Brandeis University, 415 South St. MS 015, Waltham, MA 02454-9110, USA.
出版信息
Chem Commun (Camb). 2017 Mar 2;53(19):2878-2881. doi: 10.1039/c6cc09991b.
A method for the physical attachment of folded RNA libraries to their encoding DNA is presented as a way to circumvent the reverse transcription step during systematic evolution of RNA ligands by exponential enrichment (RNA-SELEX). A DNA library is modified with one isodC base to stall T7 polymerase and a 5' "capture strand" which anneals to the nascent RNA transcript. This method is validated in a selection of RNA aptamers against human α-thrombin with dissociation constants in the low nanomolar range. This method will be useful in the discovery of RNA aptamers and ribozymes containing base modifications that make them resistant to accurate reverse transcription.
本文介绍了一种将折叠的RNA文库物理连接到其编码DNA的方法，以此规避在通过指数富集进行RNA配体的系统进化（RNA-SELEX）过程中的逆转录步骤。用一个异胞嘧啶碱基修饰DNA文库，以阻止T7聚合酶的作用，并添加一条5'“捕获链”，该链可与新生的RNA转录本退火。在针对人α-凝血酶的RNA适配体筛选中验证了该方法，所筛选出的适配体解离常数处于低纳摩尔范围。该方法将有助于发现含有碱基修饰从而使其对精确逆转录具有抗性的RNA适配体和核酶。
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