RNA-guided DNA insertion with CRISPR-associated transposases
CRISPR-Cas nucleases are powerful tools for manipulating nucleic acids; however, targeted insertion of DNA remains a challenge, as it requires host cell repair machinery. Here we characterize a CRISPR-associated transposase from cyanobacteria Scytonema hofmanni (ShCAST) that consists of Tn7-like tra...
| Main Authors: | , , , , , , |
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| Other Authors: | |
| Format: | Article |
| Language: | English |
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American Association for the Advancement of Science (AAAS)
2020
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| Online Access: | https://hdl.handle.net/1721.1/125207 |
| _version_ | 1826203895773265920 |
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| author | Strecker, Jonathan Ladha, Alim Gardner, Zachary Schmid-Burgk, Jonathan L. Makarova, Kira S. Koonin, Eugene V. Zhang, Feng |
| author2 | McGovern Institute for Brain Research at MIT |
| author_facet | McGovern Institute for Brain Research at MIT Strecker, Jonathan Ladha, Alim Gardner, Zachary Schmid-Burgk, Jonathan L. Makarova, Kira S. Koonin, Eugene V. Zhang, Feng |
| author_sort | Strecker, Jonathan |
| collection | MIT |
| description | CRISPR-Cas nucleases are powerful tools for manipulating nucleic acids; however, targeted insertion of DNA remains a challenge, as it requires host cell repair machinery. Here we characterize a CRISPR-associated transposase from cyanobacteria Scytonema hofmanni (ShCAST) that consists of Tn7-like transposase subunits and the type V-K CRISPR effector (Cas12k). ShCAST catalyzes RNA-guided DNA transposition by unidirectionally inserting segments of DNA 60 to 66 base pairs downstream of the protospacer. ShCAST integrates DNA into targeted sites in the Escherichia coli genome with frequencies of up to 80% without positive selection. This work expands our understanding of the functional diversity of CRISPR-Cas systems and establishes a paradigm for precision DNA insertion. |
| first_indexed | 2024-09-23T12:45:03Z |
| format | Article |
| id | mit-1721.1/125207 |
| institution | Massachusetts Institute of Technology |
| language | English |
| last_indexed | 2024-09-23T12:45:03Z |
| publishDate | 2020 |
| publisher | American Association for the Advancement of Science (AAAS) |
| record_format | dspace |
| spelling | mit-1721.1/1252072022-09-28T09:51:58Z RNA-guided DNA insertion with CRISPR-associated transposases Strecker, Jonathan Ladha, Alim Gardner, Zachary Schmid-Burgk, Jonathan L. Makarova, Kira S. Koonin, Eugene V. Zhang, Feng McGovern Institute for Brain Research at MIT Massachusetts Institute of Technology. Department of Brain and Cognitive Sciences Massachusetts Institute of Technology. Department of Biological Engineering CRISPR-Cas nucleases are powerful tools for manipulating nucleic acids; however, targeted insertion of DNA remains a challenge, as it requires host cell repair machinery. Here we characterize a CRISPR-associated transposase from cyanobacteria Scytonema hofmanni (ShCAST) that consists of Tn7-like transposase subunits and the type V-K CRISPR effector (Cas12k). ShCAST catalyzes RNA-guided DNA transposition by unidirectionally inserting segments of DNA 60 to 66 base pairs downstream of the protospacer. ShCAST integrates DNA into targeted sites in the Escherichia coli genome with frequencies of up to 80% without positive selection. This work expands our understanding of the functional diversity of CRISPR-Cas systems and establishes a paradigm for precision DNA insertion. National Institutes of Health (Grant 1R01-HG009761) National Institutes of Health (Grant 1R01-MH110049) National Institutes of Health (Grant 1DP1-HL141201) 2020-05-13T14:53:56Z 2020-05-13T14:53:56Z 2019-07 2019-05 2020-01-21T16:16:46Z Article http://purl.org/eprint/type/JournalArticle 0036-8075 1095-9203 https://hdl.handle.net/1721.1/125207 Strecker, Jonathan et al. "RNA-guided DNA insertion with CRISPR-associated transposases." Science 365, 6448 (July 2019): 48-53 © 2019 The Authors en http://dx.doi.org/10.1126/science.aax9181 Science Creative Commons Attribution-Noncommercial-Share Alike http://creativecommons.org/licenses/by-nc-sa/4.0/ application/pdf American Association for the Advancement of Science (AAAS) PMC |
| spellingShingle | Strecker, Jonathan Ladha, Alim Gardner, Zachary Schmid-Burgk, Jonathan L. Makarova, Kira S. Koonin, Eugene V. Zhang, Feng RNA-guided DNA insertion with CRISPR-associated transposases |
| title | RNA-guided DNA insertion with CRISPR-associated transposases |
| title_full | RNA-guided DNA insertion with CRISPR-associated transposases |
| title_fullStr | RNA-guided DNA insertion with CRISPR-associated transposases |
| title_full_unstemmed | RNA-guided DNA insertion with CRISPR-associated transposases |
| title_short | RNA-guided DNA insertion with CRISPR-associated transposases |
| title_sort | rna guided dna insertion with crispr associated transposases |
| url | https://hdl.handle.net/1721.1/125207 |
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