PCR with an expanded genetic alphabet.
PCR with an expanded genetic alphabet.
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DOI：
10.1021/ja906186f
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发表时间：
2009-10-21
影响因子：
15
通讯作者：
Romesberg, Floyd E.
中科院分区：
文献类型：
--
作者：
Malyshev, Denis A.;Seo, Young Jun;Ordoukhanian, Phillip;Romesberg, Floyd E.
用第三个碱基对扩展遗传字母表将为具有扩展遗传密码的半合成生物体奠定基础，并且也具有直接的体外应用。以前，d5 SICS和dNaM或dMMO 2之间形成的非天然碱基对被证明在稳态条件下被DNA聚合酶很好地复制，并且也被T7 RNA聚合酶在任一方向上有效地转录。我们现在证明，含有d5 SICS-dNaM或d5 SICS-dMMO 2非天然碱基对的DNA可以以接近完全天然DNA的纯度和效率进行PCR扩增。这些结果进一步证明，功能性非天然碱基对的决定簇可以被设计成主要疏水的核碱基，与天然嘌呤或嘧啶没有结构相似性。重要的是，结果表明，非天然碱基对可能在扩展的遗传字母表中发挥作用，并使许多体外应用成为可能。
Expansion of the genetic alphabet with a third base pair would lay the foundation of a semi-synthetic organism with an expanded genetic code and also have immediate in vitro applications. Previously, the unnatural base pairs formed between d5SICS and either dNaM or dMMO2 were shown to be well replicated by DNA polymerases under steady-state conditions and also transcribed by T7 RNA polymerase efficiently in either direction. We now demonstrate that DNA containing either the d5SICS-dNaM or d5SICS-dMMO2 unnatural base pair may be PCR amplified with fidelities and efficiencies that approach those of fully natural DNA. These results further demonstrate that the determinants of a functional unnatural base pair may be designed into predominantly hydrophobic nucleobases with no structural similarity to the natural purines or pyrimidines. Importantly, the results reveal that the unnatural base pairs may function within an expanded genetic alphabet and make possible many in vitro applications.
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