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Assembly of a biocompatible triazole-linked gene by one-pot click-DNA ligation.
| Content Provider | Semantic Scholar |
|---|---|
| Author | Kukwikila, M. N. Gale, Nittaya El-Sagheer, Afaf Helmy Brown, Tom Tavassoli, Ali |
| Copyright Year | 2017 |
| Abstract | The chemical synthesis of oligonucleotides and their enzyme-mediated assembly into genes and genomes has significantly advanced multiple scientific disciplines. However, these approaches are not without their shortcomings; enzymatic amplification and ligation of oligonucleotides into genes and genomes makes automation challenging, and site-specific incorporation of epigenetic information and/or modified bases into large constructs is not feasible. Here we present a fully chemical one-pot method for the assembly of oligonucleotides into a gene by click-DNA ligation. We synthesize the 335 base-pair gene that encodes the green fluorescent protein iLOV from ten functionalized oligonucleotides that contain 5'-azide and 3'-alkyne units. The resulting click-linked iLOV gene contains eight triazoles at the sites of chemical ligation, and yet is fully biocompatible; it is replicated by DNA polymerases in vitro and encodes a functional iLOV protein in Escherichia coli. We demonstrate the power and potential of our one-pot gene-assembly method by preparing an epigenetically modified variant of the iLOV gene. |
| Starting Page | 1089 |
| Ending Page | 1098 |
| Page Count | 10 |
| File Format | PDF HTM / HTML |
| DOI | 10.1038/nchem.2850 |
| PubMed reference number | 29064492 |
| Journal | Medline |
| Volume Number | 9 |
| Issue Number | 11 |
| Alternate Webpage(s) | https://media.nature.com/original/nature-assets/nchem/journal/v9/n11/extref/nchem.2850-s1.pdf |
| Alternate Webpage(s) | https://doi.org/10.1038/nchem.2850 |
| Journal | Nature chemistry |
| Language | English |
| Access Restriction | Open |
| Content Type | Text |
| Resource Type | Article |
