Evaluation of Virus-Free Manufacture of Recombinant Proteins Using CRISPR-Mediated Gene Disruption in Baculovirus-Infected Insect Cells
The manufacture and downstream processing of virus-like particles (VLPs) using the baculovirus expression vector system (BEVS) is complicated by the presence of large concentrations of baculovirus particles, which are similar in size and density to VLPs, and consequently are difficult to separate. T...
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| Format: | Article |
| Language: | English |
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MDPI AG
2023-01-01
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| Series: | Vaccines |
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| Online Access: | https://www.mdpi.com/2076-393X/11/2/225 |
| _version_ | 1797617905192402944 |
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| author | Mark R. Bruder Marc G. Aucoin |
| author_facet | Mark R. Bruder Marc G. Aucoin |
| author_sort | Mark R. Bruder |
| collection | DOAJ |
| description | The manufacture and downstream processing of virus-like particles (VLPs) using the baculovirus expression vector system (BEVS) is complicated by the presence of large concentrations of baculovirus particles, which are similar in size and density to VLPs, and consequently are difficult to separate. To reduce the burden of downstream processing, CRISPR-Cas9 technology was used to introduce insertion-deletion (indel) mutations within the <i>Autographa californica</i> multiple nucleopolyhedrovirus (<i>Ac</i>MNPV) <i>gp64</i> open reading frame, which encodes the major envelope protein of <i>Ac</i>MNPV. After comfirming the site-specific targeting of <i>gp64</i> leading to reduced budded virus (BV) release, the <i>gag</i> gene of human immunodeficiency virus type 1 was expressed to produce Gag VLPs. This approach was effective for producing VLPs using the BEVS whilst simultaneously obstructing BV release. |
| first_indexed | 2024-03-11T08:02:37Z |
| format | Article |
| id | doaj.art-37d9bb59e2414f4783e7a085dee8805e |
| institution | Directory Open Access Journal |
| issn | 2076-393X |
| language | English |
| last_indexed | 2024-03-11T08:02:37Z |
| publishDate | 2023-01-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Vaccines |
| spelling | doaj.art-37d9bb59e2414f4783e7a085dee8805e2023-11-16T23:41:29ZengMDPI AGVaccines2076-393X2023-01-0111222510.3390/vaccines11020225Evaluation of Virus-Free Manufacture of Recombinant Proteins Using CRISPR-Mediated Gene Disruption in Baculovirus-Infected Insect CellsMark R. Bruder0Marc G. Aucoin1Department of Chemical Engineering, University of Waterloo, Waterloo, ON N2L 3G1, CanadaDepartment of Chemical Engineering, University of Waterloo, Waterloo, ON N2L 3G1, CanadaThe manufacture and downstream processing of virus-like particles (VLPs) using the baculovirus expression vector system (BEVS) is complicated by the presence of large concentrations of baculovirus particles, which are similar in size and density to VLPs, and consequently are difficult to separate. To reduce the burden of downstream processing, CRISPR-Cas9 technology was used to introduce insertion-deletion (indel) mutations within the <i>Autographa californica</i> multiple nucleopolyhedrovirus (<i>Ac</i>MNPV) <i>gp64</i> open reading frame, which encodes the major envelope protein of <i>Ac</i>MNPV. After comfirming the site-specific targeting of <i>gp64</i> leading to reduced budded virus (BV) release, the <i>gag</i> gene of human immunodeficiency virus type 1 was expressed to produce Gag VLPs. This approach was effective for producing VLPs using the BEVS whilst simultaneously obstructing BV release.https://www.mdpi.com/2076-393X/11/2/225baculovirusinsect cellCRISPRvirus-like particleVLPBEVS |
| spellingShingle | Mark R. Bruder Marc G. Aucoin Evaluation of Virus-Free Manufacture of Recombinant Proteins Using CRISPR-Mediated Gene Disruption in Baculovirus-Infected Insect Cells Vaccines baculovirus insect cell CRISPR virus-like particle VLP BEVS |
| title | Evaluation of Virus-Free Manufacture of Recombinant Proteins Using CRISPR-Mediated Gene Disruption in Baculovirus-Infected Insect Cells |
| title_full | Evaluation of Virus-Free Manufacture of Recombinant Proteins Using CRISPR-Mediated Gene Disruption in Baculovirus-Infected Insect Cells |
| title_fullStr | Evaluation of Virus-Free Manufacture of Recombinant Proteins Using CRISPR-Mediated Gene Disruption in Baculovirus-Infected Insect Cells |
| title_full_unstemmed | Evaluation of Virus-Free Manufacture of Recombinant Proteins Using CRISPR-Mediated Gene Disruption in Baculovirus-Infected Insect Cells |
| title_short | Evaluation of Virus-Free Manufacture of Recombinant Proteins Using CRISPR-Mediated Gene Disruption in Baculovirus-Infected Insect Cells |
| title_sort | evaluation of virus free manufacture of recombinant proteins using crispr mediated gene disruption in baculovirus infected insect cells |
| topic | baculovirus insect cell CRISPR virus-like particle VLP BEVS |
| url | https://www.mdpi.com/2076-393X/11/2/225 |
| work_keys_str_mv | AT markrbruder evaluationofvirusfreemanufactureofrecombinantproteinsusingcrisprmediatedgenedisruptioninbaculovirusinfectedinsectcells AT marcgaucoin evaluationofvirusfreemanufactureofrecombinantproteinsusingcrisprmediatedgenedisruptioninbaculovirusinfectedinsectcells |