Sequence specific detection of DNA using nicking endonuclease signal amplification (NESA).
Sequence specific detection of DNA using nicking endonuclease signal amplification (NESA).
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DOI：
10.1093/nar/gkm654
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发表时间：
2007
影响因子：
14.9
通讯作者：
Danielsen M
中科院分区：
文献类型：
--
作者：
Kiesling T;Cox K;Davidson EA;Dretchen K;Grater G;Hibbard S;Lasken RS;Leshin J;Skowronski E;Danielsen M
我们开发了一种识别特定单链或双链 DNA 序列的新方法，称为切口核酸内切酶信号放大 (NESA)。探针和目标 DNA 退火以创建限制位点，该限制位点可被链特异性核酸内切酶识别，将探针切割成两段，而目标 DNA 保持完整。然后，目标 DNA 可以作为新探针的模板，并重复杂交、切割和解离过程。激光诱导荧光与毛细管电泳结合用于测量探针裂解产物。反应迅速；在理想条件下，探针在一分钟内完全裂解。该反应是特异性的，因为它需要寡核苷酸和模板在限制性位点处完全互补，并且总体上具有足够的互补性以允许杂交。我们表明，枯草芽孢杆菌和炭疽芽孢杆菌基因组 DNA 都可以被检测到，并与其他芽孢杆菌属物种的 DNA 进行特异性区分。当与多重置换扩增相结合时，可以检测小于 30 cfu 的单拷贝靶标。每当需要检测特定 DNA 序列时，都应适用此方法。其他应用包括 SNP 分析和基因分型。该反应本身就很容易进行多重反应，并且适合自动化。
We have developed a new method for identifying specific single- or double-stranded DNA sequences called nicking endonuclease signal amplification (NESA). A probe and target DNA anneal to create a restriction site that is recognized by a strand-specific endonuclease that cleaves the probe into two pieces leaving the target DNA intact. The target DNA can then act as a template for fresh probe and the process of hybridization, cleavage and dissociation repeats. Laser-induced fluorescence coupled with capillary electrophoresis was used to measure the probe cleavage products. The reaction is rapid; full cleavage of probe occurs within one minute under ideal conditions. The reaction is specific since it requires complete complementarity between the oligonucleotide and the template at the restriction site and sufficient complementarity overall to allow hybridization. We show that both Bacillus subtilis and B. anthracis genomic DNA can be detected and specifically differentiated from DNA of other Bacillus species. When combined with multiple displacement amplification, detection of a single copy target from less than 30 cfu is possible. This method should be applicable whenever there is a requirement to detect a specific DNA sequence. Other applications include SNP analysis and genotyping. The reaction is inherently simple to multiplex and is amenable to automation.
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影响因子：
5.1
作者：
Gonzalez, JM;Portillo, MC;Saiz-Jimenez, C
通讯作者：
Saiz-Jimenez, C
影响因子：
7
作者：
Hosono, S;Faruqi, AF;Lasken, RS
通讯作者：
Lasken, RS
影响因子：
3.5
作者：
Kim, YG;Shi, YG;Chandrasegaran, S
通讯作者：
Chandrasegaran, S
影响因子：
3.3
作者：
Burton, JE;Oshota, J;Silman, NJ
通讯作者：
Silman, NJ
影响因子：
14.9
作者：
NASRI, M;THOMAS, D
通讯作者：
THOMAS, D