High-throughput functional variant screens via in vivo production of single-stranded DNA
<jats:title>Significance</jats:title> <jats:p>We report a methodology for the pooled construction of mutants bearing precise genomic sequence variations and multiplex phenotypic characterization of these mutants using next-generation sequencing (NGS). Unlike existing tec...
| Main Authors: | , , , , , , , |
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| Other Authors: | |
| Format: | Article |
| Language: | English |
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Proceedings of the National Academy of Sciences
2022
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| Online Access: | https://hdl.handle.net/1721.1/143760 |
| _version_ | 1826209916114698240 |
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| author | Schubert, Max G Goodman, Daniel B Wannier, Timothy M Kaur, Divjot Farzadfard, Fahim Lu, Timothy K Shipman, Seth L Church, George M |
| author2 | Massachusetts Institute of Technology. Research Laboratory of Electronics |
| author_facet | Massachusetts Institute of Technology. Research Laboratory of Electronics Schubert, Max G Goodman, Daniel B Wannier, Timothy M Kaur, Divjot Farzadfard, Fahim Lu, Timothy K Shipman, Seth L Church, George M |
| author_sort | Schubert, Max G |
| collection | MIT |
| description | <jats:title>Significance</jats:title>
<jats:p>We report a methodology for the pooled construction of mutants bearing precise genomic sequence variations and multiplex phenotypic characterization of these mutants using next-generation sequencing (NGS). Unlike existing techniques depending on CRISPR-Cas–directed genomic breaks for genome editing, this strategy instead uses single-stranded DNA produced by a retron element for recombineering. This enables libraries of millions of elements to be constructed and offers relaxed design constraints which permit natural DNA or random variation to be used as inputs.</jats:p> |
| first_indexed | 2024-09-23T14:35:16Z |
| format | Article |
| id | mit-1721.1/143760 |
| institution | Massachusetts Institute of Technology |
| language | English |
| last_indexed | 2024-09-23T14:35:16Z |
| publishDate | 2022 |
| publisher | Proceedings of the National Academy of Sciences |
| record_format | dspace |
| spelling | mit-1721.1/1437602023-04-13T20:38:44Z High-throughput functional variant screens via in vivo production of single-stranded DNA Schubert, Max G Goodman, Daniel B Wannier, Timothy M Kaur, Divjot Farzadfard, Fahim Lu, Timothy K Shipman, Seth L Church, George M Massachusetts Institute of Technology. Research Laboratory of Electronics <jats:title>Significance</jats:title> <jats:p>We report a methodology for the pooled construction of mutants bearing precise genomic sequence variations and multiplex phenotypic characterization of these mutants using next-generation sequencing (NGS). Unlike existing techniques depending on CRISPR-Cas–directed genomic breaks for genome editing, this strategy instead uses single-stranded DNA produced by a retron element for recombineering. This enables libraries of millions of elements to be constructed and offers relaxed design constraints which permit natural DNA or random variation to be used as inputs.</jats:p> 2022-07-15T13:29:12Z 2022-07-15T13:29:12Z 2021 2022-07-15T13:08:41Z Article http://purl.org/eprint/type/JournalArticle https://hdl.handle.net/1721.1/143760 Schubert, Max G, Goodman, Daniel B, Wannier, Timothy M, Kaur, Divjot, Farzadfard, Fahim et al. 2021. "High-throughput functional variant screens via in vivo production of single-stranded DNA." Proceedings of the National Academy of Sciences of the United States of America, 118 (18). en 10.1073/PNAS.2018181118 Proceedings of the National Academy of Sciences of the United States of America Article is made available in accordance with the publisher's policy and may be subject to US copyright law. Please refer to the publisher's site for terms of use. application/pdf Proceedings of the National Academy of Sciences PNAS |
| spellingShingle | Schubert, Max G Goodman, Daniel B Wannier, Timothy M Kaur, Divjot Farzadfard, Fahim Lu, Timothy K Shipman, Seth L Church, George M High-throughput functional variant screens via in vivo production of single-stranded DNA |
| title | High-throughput functional variant screens via in vivo production of single-stranded DNA |
| title_full | High-throughput functional variant screens via in vivo production of single-stranded DNA |
| title_fullStr | High-throughput functional variant screens via in vivo production of single-stranded DNA |
| title_full_unstemmed | High-throughput functional variant screens via in vivo production of single-stranded DNA |
| title_short | High-throughput functional variant screens via in vivo production of single-stranded DNA |
| title_sort | high throughput functional variant screens via in vivo production of single stranded dna |
| url | https://hdl.handle.net/1721.1/143760 |
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