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| Indexado |
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| DOI | 10.1111/NPH.15625 | ||||
| Año | 2019 | ||||
| Tipo | artículo de investigación |
Citas Totales
Autores Afiliación Chile
Instituciones Chile
% Participación
Internacional
Autores
Afiliación Extranjera
Instituciones
Extranjeras
High-efficiency methods for DNA assembly have enabled the routine assembly of synthetic DNAs of increased size and complexity. However, these techniques require customization, elaborate vector sets or serial manipulations for the different stages of assembly. We have developed Loop assembly based on a recursive approach to DNA fabrication. The system makes use of two Type IIS restriction endonucleases and corresponding vector sets for efficient and parallel assembly of large DNA circuits. Standardized level 0 parts can be assembled into circuits containing 1, 4, 16 or more genes by looping between the two vector sets. The vectors also contain modular sites for hybrid assembly using sequence overlap methods. Loop assembly enables efficient and versatile DNA fabrication for plant transformation. We show the construction of plasmids up to 16 genes and 38 kb with high efficiency (> 80%). We have characterized Loop assembly on over 200 different DNA constructs and validated the fidelity of the method by high-throughput Illumina plasmid sequencing. Our method provides a simple generalized solution for DNA construction with standardized parts. The cloning system is provided under an OpenMTA license for unrestricted sharing and open access.
| Ord. | Autor | Género | Institución - País |
|---|---|---|---|
| 1 | Pollak, Bernardo | Hombre |
UNIV CAMBRIDGE - Reino Unido
University of Cambridge - Reino Unido |
| 2 | Cerda, Ariel | Hombre |
Pontificia Universidad Católica de Chile - Chile
Núcleo Milenio en Biología Sintética y Biología de Sistemas Vegetales - Chile Fondo de Desarrollo de Áreas Prioritarias - Chile |
| 3 | Delmans, Mihails | Hombre |
UNIV CAMBRIDGE - Reino Unido
University of Cambridge - Reino Unido |
| 4 | Alamos, Simon | Hombre |
UNIV CALIF BERKELEY - Estados Unidos
Department of Plant & Microbial Biology - Estados Unidos |
| 5 | MOYANO-YUGOVIC, TOMAS CUSTODIO | Hombre |
Pontificia Universidad Católica de Chile - Chile
Núcleo Milenio en Biología Sintética y Biología de Sistemas Vegetales - Chile Fondo de Desarrollo de Áreas Prioritarias - Chile |
| 6 | West, Anthony | Hombre |
Earlham Inst - Reino Unido
Earlham Institute - Reino Unido |
| 7 | GUTIERREZ-ILABACA, RODRIGO ANTONIO | Hombre |
Pontificia Universidad Católica de Chile - Chile
Núcleo Milenio en Biología Sintética y Biología de Sistemas Vegetales - Chile Fondo de Desarrollo de Áreas Prioritarias - Chile |
| 8 | Patron, Nicola J. | Mujer |
Earlham Inst - Reino Unido
Earlham Institute - Reino Unido |
| 9 | Federici, Fernan | - |
UNIV CAMBRIDGE - Reino Unido
Pontificia Universidad Católica de Chile - Chile Núcleo Milenio en Biología Sintética y Biología de Sistemas Vegetales - Chile University of Cambridge - Reino Unido Fondo de Desarrollo de Áreas Prioritarias - Chile |
| 10 | Haseloff, Jim | - |
UNIV CAMBRIDGE - Reino Unido
University of Cambridge - Reino Unido |
| Fuente |
|---|
| CONICYT FONDECYT |
| Biotechnology and Biological Sciences Research Council |
| Becas Chile |
| Harvard University |
| Biotechnology and Biological Sciences Research Council (BBSRC) |
| CONICYT Fondecyt Iniciacion |
| Engineering and Physical Sciences Research Council |
| Center for Genome Regulation |
| Fondos de Desarrollo de la Astronomía Nacional |
| Instituto Milenio iBio - Iniciativa Cientifica Milenio MINECON |
| Fondo de Desarrollo de Areas Prioritarias (FONDAP) Center for Genome Regulation |
| Cambridge Commonwealth Trust |
| Fondo de Desarrollo de Áreas Prioritarias |
| BBSRC National Capability in Genomics at the Earlham Institute |
| Cambridge Trust |
| University of Cambridge BBSRC DTP programme |
| Gates Cambridge Trust |
| Instituto Milenio iBio – Iniciativa Científica Milenio MINECON |
| CONICYT Fon-decyt Iniciación |
| University of Cambridge BBSRC |
| CONICYT Fon-decyt |
| Georg-August-Universität Göttingen |
| Earlham Institute |
| Instituto de Biología Molecular y Celular de Plantas |
| Teva Vernaux |
| BioBricks Foundation |
| Agradecimiento |
|---|
| We would like to acknowledge Diego Orzaez (Instituto de Biologia Molecular y Celular de Plantas) for helpful suggestions and advice regarding SapI and Type IIS restriction enzymes, Oleg Raitskin (Earlham Institute) for advice on Type IIS assembly protocols, H. Ghareeb (Gottingen University), T. W. J. Gadella and J. Goedhart (both University of Amsterdam) for providing mTurquoise2 DNA, Jen Sheen (Harvard University) for providing TCS promoter DNA, Teva Vernaux (Ecole Normale Superieure de Lyon) for Venus N7 DNA, Jeff Lichtman (Harvard University) for mTagBFP2 DNA, and Susana Sauret-Gueto, Eftychis Frangedakis and Marta Tomaselli (all University of Cambridge) for troubleshooting SapI reactions. We thank Linda Kahl and Drew Endy (BioBricks Foundation) for discussions on the implementation of the OpenMTA. Support for the authors was provided by Becas Chile and the Cambridge Trust (to BP), University of Cambridge BBSRC DTP programme (to MD) and the Biotechnology and Biological Sciences Research Council (BBSRC) and Engineering and Physical Sciences Research Council (OpenPlant Grant no. BB/L014130/1) (to NJP, FF and JH). Laboratory automation, next-generation sequencing and library construction was delivered via the BBSRC National Capability in Genomics (BB/CCG1720/1) at the Earlham Institute. FF acknowledges funding from CONICYT Fondecyt Iniciacion 11140776. FF and RAG acknowledge funding from the Fondo de Desarrollo de Areas Prioritarias (FONDAP) Center for Genome Regulation (15090007) and Instituto Milenio iBio - Iniciativa Cientifica Milenio MINECON. The authors declare that they have no competing financial interests. |
| We would like to acknowledge Diego Orzaez (Instituto de Biología Molecular y Celular de Plantas) for helpful suggestions and advice regarding SapI and Type IIS restriction enzymes, Oleg Raitskin (Earlham Institute) for advice on Type IIS assembly protocols, H. Ghareeb (G€ottingen University), T. W. J. Gadella and J. Goedhart (both University of Amsterdam) for providing mTurquoise2 DNA, Jen Sheen (Harvard University) for providing TCS promoter DNA, Teva Vernaux (Ecole Normale Supérieure de Lyon) for Venus N7 DNA, Jeff Lichtman (Harvard University) for mTagBFP2 DNA, and Susana Sauret-Gueto, Eftychis Frangedakis and Marta Tomaselli (all University of Cambridge) for troubleshooting SapI reactions. We thank Linda Kahl and Drew Endy (BioBricks Foundation) for discussions on the implementation of the OpenMTA. Support for the authors was provided by Becas Chile and the Cambridge Trust (to BP), University of Cambridge BBSRC DTP programme (to MD) and the Biotechnology and Biological Sciences Research Council (BBSRC) and Engineering and Physical Sciences Research Council (OpenPlant Grant no. BB/L014130/1) (to NJP, FF and JH). Laboratory automation, next-generation sequencing and library construction was delivered via the BBSRC National Capability in Genomics (BB/CCG1720/1) at the Earl-ham Institute. FF acknowledges funding from CONICYT Fon-decyt Iniciación 11140776. FF and RAG acknowledge funding from the Fondo de Desarrollo de Areas Prioritarias (FONDAP) Center for Genome Regulation (15090007) and Instituto Milenio iBio – Iniciativa Científica Milenio MINECON. The authors declare that they have no competing financial interests. |