A new class of homogeneous nucleic acid probes based on specific displacement hybridization
A new class of homogeneous nucleic acid probes based on specific displacement hybridization
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DOI：
10.1093/nar/30.2.e5
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发表时间：
2002-01-15
影响因子：
14.9
通讯作者：
Liang, JX
中科院分区：
文献类型：
--
作者：
Li, QQ;Luan, GY;Liang, JX
我们已经开发了一类新的探针，用于同质核酸检测的基础上提出的置换杂交。我们的探针由两个互补的不同长度的寡脱氧核糖核苷酸标记的荧光团和猝灭剂在双链体中的紧密接近。探针本身被淬灭，但它们在与靶标置换杂交后变得发荧光。这些探针显示完全匹配的靶和单核苷酸错配靶之间的完全区分。双链探针与相应的线性探针的比较证实，互补链的存在显着增强其特异性。使用四个这样的探针标记不同颜色的荧光团，每个设计识别不同的目标，我们已经证明，多个目标可以区分在同一个解决方案，即使他们彼此之间的差异少到一个核苷酸。在实时核酸扩增中使用双链探针作为探针或引物。除了具有极高的特异性和灵活性外，这类新的探针设计和合成简单，成本低，灵敏度高，并且可用于各种标记。这类探针应该在核酸杂交的高特异性相关的各种领域中找到应用。
We have developed a new class of probes for homogeneous nucleic acid detection based on the proposed displacement hybridization. Our probes consist of two complementary oligodeoxyribonucleotides of different length labeled with a fluorophore and a quencher in close proximity in the duplex. The probes on their own are quenched, but they become fluorescent upon displacement hybridization with the target. These probes display complete discrimination between a perfectly matched target and single nucleotide mismatch targets. A comparison of double-stranded probes with corresponding linear probes confirms that the presence of the complementary strand significantly enhances their specificity. Using four such probes labeled with different color fluorophores, each designed to recognize a different target, we have demonstrated that multiple targets can be distinguished in the same solution, even if they differ from one another by as little as a single nucleotide. Double-stranded probes were used in real-time nucleic acid amplifications as either probes or as primers. In addition to its extreme specificity and flexibility, the new class of probes is simple to design and synthesize, has low cost and high sensitivity and is accessible to a wide range of labels. This class of probes should find applications in a variety of areas wherever high specificity of nucleic acid hybridization is relevant.