Gibson assembly interposition improves amplification efficiency of long DNA and multifragment overlap extension PCR
For difficult overlap extension PCR, a Gibson assembly process was inserted between the two PCR rounds to facilitate the formation of complete gene templates at a moderate temperature. That is, after amplifying each DNA fragment, they were preluded by a Gibson assembly process in equal proportion. T...
| Veröffentlicht in: | BioTechniques. - 1993. - 74(2023), 6 vom: 31. Juni, Seite 286-292 |
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| Format: | Online-Aufsatz |
| Sprache: | English |
| Veröffentlicht: |
2023
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| Zugriff auf das übergeordnete Werk: | BioTechniques |
| Schlagworte: | Journal Article Research Support, Non-U.S. Gov't Gibson assembly gene fusion overlap extension PCR retinoblastoma gene site-directed mutagenesis DNA 9007-49-2 |
| Zusammenfassung: | For difficult overlap extension PCR, a Gibson assembly process was inserted between the two PCR rounds to facilitate the formation of complete gene templates at a moderate temperature. That is, after amplifying each DNA fragment, they were preluded by a Gibson assembly process in equal proportion. Then, the assembled mixture was used as a template for the second PCR round. This idea was tested and verified by taking the cloning example of a single and a double site mutation of the retinoblastoma gene. This scheme associates overlap extension PCR with Gibson assembly exquisitely, significantly improving gene amplification efficiency, particularly in the fusion of long genes and multifragments using overlap extension PCR |
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| Beschreibung: | Date Completed 26.07.2023 Date Revised 26.07.2023 published: Print-Electronic Citation Status MEDLINE |
| ISSN: | 1940-9818 |
| DOI: | 10.2144/btn-2023-0012 |