| Autor: |
Palluk, S, Arlow, DH, De Rond, T, Barthel, S, Kang, JS, Bector, R, Baghdassarian, HM, Truong, AN, Kim, PW, Singh, AK, Hillson, NJ, Keasling, JD |
| Jazyk: |
angličtina |
| Rok vydání: |
2018 |
| Předmět: |
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| Zdroj: |
Palluk, S; Arlow, DH; De Rond, T; Barthel, S; Kang, JS; Bector, R; et al.(2018). De novo DNA synthesis using polymerasenucleotide conjugates. Nature Biotechnology, 36(7), 645-650. doi: 10.1038/nbt.4173. UC Berkeley: Retrieved from: http://www.escholarship.org/uc/item/9d4221mc |
| Popis: |
© 2018 Nature Publishing Group. All rights reserved. Oligonucleotides are almost exclusively synthesized using the nucleoside phosphoramidite method, even though it is limited to the direct synthesis of ∼200 mers and produces hazardous waste. Here, we describe an oligonucleotide synthesis strategy that uses the template-independent polymerase terminal deoxynucleotidyl transferase (TdT). Each TdT molecule is conjugated to a single deoxyribonucleoside triphosphate (dNTP) molecule that it can incorporate into a primer. After incorporation of the tethered dNTP, the 3′ end of the primer remains covalently bound to TdT and is inaccessible to other TdT-dNTP molecules. Cleaving the linkage between TdT and the incorporated nucleotide releases the primer and allows subsequent extension. We demonstrate that TdT-dNTP conjugates can quantitatively extend a primer by a single nucleotide in 10-20 s, and that the scheme can be iterated to write a defined sequence. This approach may form the basis of an enzymatic oligonucleotide synthesizer. |
| Databáze: |
OpenAIRE |
| Externí odkaz: |
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