Capillary electrophoresis of DNA for molecular diagnostics
Capillary electrophoresis of DNA for molecular diagnostics
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DOI：
10.1002/elps.1150181003
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发表时间：
1997
期刊：
影响因子：
2.9
通讯作者：
P. Righetti;C. Gelfi
中科院分区：
文献类型：
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作者：
P. Righetti;C. Gelfi
综述了毛细管区带电泳(CZE)在筛选液体聚合物(主要是线性聚丙烯酰胺和纤维素)中的一些应用，以分析临床上相关的诊断DNA的聚合酶链式反应(PCR)产物。涉及的领域包括：人类遗传学、定量基因剂量、微生物学和病毒学、法医学和治疗性DNA(特别是反义核苷酸)。重点介绍了一些独特的新进展，例如：(I)非等规共沉淀CZE，即用于检测DNA点突变的程序升温CZE；(Ii)新型N-取代丙烯酰胺的合成，具有极高的抗碱性水解性和高亲水性。在变性梯度凝胶电泳(DGGE)领域，正如在凝胶平板上常规进行的那样，在CZE中描述了一种新的方法：双梯度DGGE。在这项技术中，沿迁移方向同时应用两个梯度：化学(或热)变性梯度，用于部分解开DNA的同/异双链；以及孔隙度梯度，用于重新压缩在更广泛的变性条件下熔化的扩散带。由此证明，除了温度梯度之外，即使在毛细管格式中也可以很容易地实现化学梯度。
A number of applications of capillary zone electrophoresis (CZE) in sieving liquid polymers (notably linear polyacrylamides and cellulose) for the analysis of polymerase chain reaction (PCR) products of clinically relevant, diagnostic DNA, are reviewed. The fields covered are: human genetics, quantitative gene dosage, microbiology and virology, forensic medicine and therapeutic DNA (notably, antisense nucleotides). Some unique, novel developments are highlighted, such as: (i) nonisocratic CZE, i.e., temperature‐programmed CZE for detection of DNA point mutations; (ii) the synthesis of novel N‐substituted acrylamides, offering extreme resistance to alkaline hydrolysis coupled to high hydrophilicity. In the field of denaturing gradient gel electrophoresis (DGGE), as routinely performed in gel slabs, a novel methodology is described in CZE: double‐gradient DGGE. In this technique, two gradients are simultaneously applied along the migration direction: a chemical (or thermal) denaturing gradient, for partially unwinding homo‐ and hetero‐duplexes of DNA, and a porosity gradient, for recompacting diffuse bands melting over a broader range of denaturing conditions. It is thus demonstrated that chemical gradients, in addition to temperature gradients, can be easily implemented even in a capillary format.