基于DNAzyme的d(GGA)重复序列折叠结构的可视检测  

G-quadruple/Hemin DNAzyme-based Visual Detection of the Folding Structure Formed by d(GGA)Trinucleotide Repeats

作　　者：游昕 贺荣荣 闫兴贺 南银杏 常天俊[1] 邴涛 YOU Xin;HE Rong-rong;YAN Xing-he;NAN Yin-xing;CHANG Tian-jun;BING Tao(Institute of Resources and Environment,Henan Polytechnic University,Jiaozuo 454003,China;Hangzhou Institute of Medicine(HIM),Chinese Academy of Sciences,Hangzhou 310022,China)

机构地区：[1]河南理工大学资源环境学院,河南焦作454003 [2]中国科学院杭州医学研究所,浙江杭州310022

出　　处：《分析测试学报》2023年第11期1495-1502,共8页Journal of Instrumental Analysis

基　　金：国家自然科学基金项目(U1704241);浙江省核酸适体与临床诊治重点实验室开放基金项目(2022ASC001)。

摘　　要：d(GGA)重复序列形成的H(G-A七分体):T(G-四分体)型“非常规”G-四链体(G4)及其二聚体是其调控c-myb原癌基因表达的关键。该类序列也在很多其他基因中存在,被认为可能具有与其折叠相关的功能,因此判别其折叠结构是其功能研究的基础。该文发现d(GGA)4与血红素可形成高活性DNAzyme,催化裸眼可视的快速显色反应,且DNAzyme活性与其结构相关,基于此提出了可视检测d(GGA)重复序列折叠结构的策略。通过研究20条含不同侧翼和重复次数不同的d(GGA)重复序列,发现16条有高DNAzyme活性的序列具有高比例的G4二聚体,4条DNAzyme活性显著降低或接近失活的序列聚集状态很少。表明DNAzyme活性与d(GGA)重复序列折叠成的G4二聚体高度相关,基于DNAzyme活性的分析方法可用于其在实际序列中折叠结构的可视检测。d(GGA)trinucleotide repeats play an important role in the regulation of the expression of c-myb proto-oncogene.Four GGA repeats have proven to adopt an“unconventional”G-quadruplex(G4)including a 3’guanine tetrad(T)stacked onto a 5’guanine-adenine heptad(H).Moreover,two d(GGA)4 form very stable inter-molecular G4 by stacking two T∶H G-quadruplexes on the heptad plane,resulting in a T∶H∶H∶T dimer.d(GGA)trinucleotide repeats have been frequently found from the genome of various organisms and suggested to have the structure-based functions.Thus,it is necessary to identify the folding structures of these sequences before studying their biological functions.In this work,d(GGA)4 was discovered can be made of highly active DNAzyme in the presence of hemin,which catalyze a rapid reaction visualized by naked eyes.This DNAzyme activity is related to its folding structure.Based on the finding,a visualization method was raised for the rapid identification of the folding structures by d(GGA)trinucleotide repeats.A total of 20 sequences with d(GGA)repeats that were flanked with different nucleotides or with various repeat numbers were investigated.16 sequences that possessed high DNAzyme activity were revealed to adopt dimer with a large proportion by gel electrophoresis,yet 4 sequences with dramatically reduced or no DNAzyme activity were found nearly no G4 or assembled fractions.These results suggest the highly reliable for using DNAzyme assay to visualization the folding structures by d(GGA)repeats in real sequences,expanding the usage of DNAzyme assay for the detection of“unconventional”G4s.

关 键 词：G-四链体 DNAZYME 血红素 核酸适体 生物传感器 可视检测 

分 类 号：O629.7[理学—有机化学] TP212.3[理学—化学]