A Baculovirus Expression Vector Derived Entirely from Non-Templated, Chemically Synthesized DNA Parts
Baculovirus expression system1s are a widely used tool in recombinant protein and biologics production. To enable the possibility of genome modifications unconstrained through low-throughput and bespoke classical genome manipulation techniques, we set out to construct a baculovirus vector (>130 k...
| Main Authors: | , , |
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| Format: | Article |
| Language: | English |
| Published: |
MDPI AG
2023-09-01
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| Series: | Viruses |
| Subjects: | |
| Online Access: | https://www.mdpi.com/1999-4915/15/10/1981 |
| _version_ | 1827719712644005888 |
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| author | Christopher Nguyen Amanda Ibe-Enwo Jeffrey Slack |
| author_facet | Christopher Nguyen Amanda Ibe-Enwo Jeffrey Slack |
| author_sort | Christopher Nguyen |
| collection | DOAJ |
| description | Baculovirus expression system1s are a widely used tool in recombinant protein and biologics production. To enable the possibility of genome modifications unconstrained through low-throughput and bespoke classical genome manipulation techniques, we set out to construct a baculovirus vector (>130 kb dsDNA) built from modular, chemically synthesized DNA parts. We constructed a synthetic version of <i>Autographa californica</i> multiple nucleopolyhedrovirus (<i>Ac</i>MNPV) through two steps of hierarchical Golden Gate assembly. Over 140 restriction endonuclease sites were removed to enable the discrimination of the synthetic genome from native baculovirus genomes. A head-to-head comparison of our modular, synthetic <i>Ac</i>MNPV genome with native baculovirus vectors showed no significant difference in baculovirus growth kinetics or recombinant adeno-associated virus production—suggesting that neither baculovirus replication nor very-late gene expression were compromised by our design or assembly method. With unprecedented control over the <i>Ac</i>MNPV genome at the single-nucleotide level, we hope to ambitiously explore novel <i>Ac</i>MNPV vectors streamlined for biologics production and development. |
| first_indexed | 2024-03-10T20:49:58Z |
| format | Article |
| id | doaj.art-7cd7b80a7ffb4dd2ad3a55a2472f5108 |
| institution | Directory Open Access Journal |
| issn | 1999-4915 |
| language | English |
| last_indexed | 2024-03-10T20:49:58Z |
| publishDate | 2023-09-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Viruses |
| spelling | doaj.art-7cd7b80a7ffb4dd2ad3a55a2472f51082023-11-19T18:26:25ZengMDPI AGViruses1999-49152023-09-011510198110.3390/v15101981A Baculovirus Expression Vector Derived Entirely from Non-Templated, Chemically Synthesized DNA PartsChristopher Nguyen0Amanda Ibe-Enwo1Jeffrey Slack2Stylus Medicine, Inc., 200 Berkeley St., Boston, MA 02116, USAVoyager Therapeutics, 64 Sidney St., Cambridge, MA 02139, USAVoyager Therapeutics, 64 Sidney St., Cambridge, MA 02139, USABaculovirus expression system1s are a widely used tool in recombinant protein and biologics production. To enable the possibility of genome modifications unconstrained through low-throughput and bespoke classical genome manipulation techniques, we set out to construct a baculovirus vector (>130 kb dsDNA) built from modular, chemically synthesized DNA parts. We constructed a synthetic version of <i>Autographa californica</i> multiple nucleopolyhedrovirus (<i>Ac</i>MNPV) through two steps of hierarchical Golden Gate assembly. Over 140 restriction endonuclease sites were removed to enable the discrimination of the synthetic genome from native baculovirus genomes. A head-to-head comparison of our modular, synthetic <i>Ac</i>MNPV genome with native baculovirus vectors showed no significant difference in baculovirus growth kinetics or recombinant adeno-associated virus production—suggesting that neither baculovirus replication nor very-late gene expression were compromised by our design or assembly method. With unprecedented control over the <i>Ac</i>MNPV genome at the single-nucleotide level, we hope to ambitiously explore novel <i>Ac</i>MNPV vectors streamlined for biologics production and development.https://www.mdpi.com/1999-4915/15/10/1981baculovirus expression vectorsynthetic biologyrAAVGolden Gate assemblysynthetic virus genome |
| spellingShingle | Christopher Nguyen Amanda Ibe-Enwo Jeffrey Slack A Baculovirus Expression Vector Derived Entirely from Non-Templated, Chemically Synthesized DNA Parts Viruses baculovirus expression vector synthetic biology rAAV Golden Gate assembly synthetic virus genome |
| title | A Baculovirus Expression Vector Derived Entirely from Non-Templated, Chemically Synthesized DNA Parts |
| title_full | A Baculovirus Expression Vector Derived Entirely from Non-Templated, Chemically Synthesized DNA Parts |
| title_fullStr | A Baculovirus Expression Vector Derived Entirely from Non-Templated, Chemically Synthesized DNA Parts |
| title_full_unstemmed | A Baculovirus Expression Vector Derived Entirely from Non-Templated, Chemically Synthesized DNA Parts |
| title_short | A Baculovirus Expression Vector Derived Entirely from Non-Templated, Chemically Synthesized DNA Parts |
| title_sort | baculovirus expression vector derived entirely from non templated chemically synthesized dna parts |
| topic | baculovirus expression vector synthetic biology rAAV Golden Gate assembly synthetic virus genome |
| url | https://www.mdpi.com/1999-4915/15/10/1981 |
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