Streamlining recombination-mediated genetic engineering by validating three neutral integration sites in Synechococcus sp. PCC 7002
Abstract Background Synechococcus sp. PCC 7002 (henceforth Synechococcus) is developing into a powerful synthetic biology chassis. In order to streamline the integration of genes into the Synechococcus chromosome, validation of neutral integration sites with optimization of the DNA transformation pr...
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| Format: | Article |
| Language: | English |
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BMC
2017-06-01
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| Series: | Journal of Biological Engineering |
| Subjects: | |
| Online Access: | http://link.springer.com/article/10.1186/s13036-017-0061-8 |
| _version_ | 1798045395700416512 |
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| author | Anne Ilse Maria Vogel Rahmi Lale Martin Frank Hohmann-Marriott |
| author_facet | Anne Ilse Maria Vogel Rahmi Lale Martin Frank Hohmann-Marriott |
| author_sort | Anne Ilse Maria Vogel |
| collection | DOAJ |
| description | Abstract Background Synechococcus sp. PCC 7002 (henceforth Synechococcus) is developing into a powerful synthetic biology chassis. In order to streamline the integration of genes into the Synechococcus chromosome, validation of neutral integration sites with optimization of the DNA transformation protocol parameters is necessary. Availability of BioBrick-compatible integration modules is desirable to further simplifying chromosomal integrations. Results We designed three BioBrick-compatible genetic modules, each targeting a separate neutral integration site, A2842, A0935, and A0159, with varying length of homologous region, spanning from 100 to 800 nt. The performance of the different modules for achieving DNA integration were tested. Our results demonstrate that 100 nt homologous regions are sufficient for inserting a 1 kb DNA fragment into the Synechococcus chromosome. By adapting a transformation protocol from a related cyanobacterium, we shortened the transformation procedure for Synechococcus significantly. Conclusions The optimized transformation protocol reported in this study provides an efficient way to perform genetic engineering in Synechococcus. We demonstrated that homologous regions of 100 nt are sufficient for inserting a 1 kb DNA fragment into the three tested neutral integration sites. Integration at A2842, A0935 and A0159 results in only a minimal fitness cost for the chassis. This study contributes to developing Synechococcus as the prominent chassis for future synthetic biology applications. |
| first_indexed | 2024-04-11T23:20:11Z |
| format | Article |
| id | doaj.art-a54b82a613824f7abb027d43f591fb6c |
| institution | Directory Open Access Journal |
| issn | 1754-1611 |
| language | English |
| last_indexed | 2024-04-11T23:20:11Z |
| publishDate | 2017-06-01 |
| publisher | BMC |
| record_format | Article |
| series | Journal of Biological Engineering |
| spelling | doaj.art-a54b82a613824f7abb027d43f591fb6c2022-12-22T03:57:28ZengBMCJournal of Biological Engineering1754-16112017-06-011111910.1186/s13036-017-0061-8Streamlining recombination-mediated genetic engineering by validating three neutral integration sites in Synechococcus sp. PCC 7002Anne Ilse Maria Vogel0Rahmi Lale1Martin Frank Hohmann-Marriott2Department of Biotechnology, PhotoSynLab, NTNU, Norwegian University of Science and TechnologyDepartment of Biotechnology, PhotoSynLab, NTNU, Norwegian University of Science and TechnologyDepartment of Biotechnology, PhotoSynLab, NTNU, Norwegian University of Science and TechnologyAbstract Background Synechococcus sp. PCC 7002 (henceforth Synechococcus) is developing into a powerful synthetic biology chassis. In order to streamline the integration of genes into the Synechococcus chromosome, validation of neutral integration sites with optimization of the DNA transformation protocol parameters is necessary. Availability of BioBrick-compatible integration modules is desirable to further simplifying chromosomal integrations. Results We designed three BioBrick-compatible genetic modules, each targeting a separate neutral integration site, A2842, A0935, and A0159, with varying length of homologous region, spanning from 100 to 800 nt. The performance of the different modules for achieving DNA integration were tested. Our results demonstrate that 100 nt homologous regions are sufficient for inserting a 1 kb DNA fragment into the Synechococcus chromosome. By adapting a transformation protocol from a related cyanobacterium, we shortened the transformation procedure for Synechococcus significantly. Conclusions The optimized transformation protocol reported in this study provides an efficient way to perform genetic engineering in Synechococcus. We demonstrated that homologous regions of 100 nt are sufficient for inserting a 1 kb DNA fragment into the three tested neutral integration sites. Integration at A2842, A0935 and A0159 results in only a minimal fitness cost for the chassis. This study contributes to developing Synechococcus as the prominent chassis for future synthetic biology applications.http://link.springer.com/article/10.1186/s13036-017-0061-8Synechococcus sp. PCC7002Neutral integration sitesGenetic engineeringTransformationBioBrickCyanobacteria |
| spellingShingle | Anne Ilse Maria Vogel Rahmi Lale Martin Frank Hohmann-Marriott Streamlining recombination-mediated genetic engineering by validating three neutral integration sites in Synechococcus sp. PCC 7002 Journal of Biological Engineering Synechococcus sp. PCC7002 Neutral integration sites Genetic engineering Transformation BioBrick Cyanobacteria |
| title | Streamlining recombination-mediated genetic engineering by validating three neutral integration sites in Synechococcus sp. PCC 7002 |
| title_full | Streamlining recombination-mediated genetic engineering by validating three neutral integration sites in Synechococcus sp. PCC 7002 |
| title_fullStr | Streamlining recombination-mediated genetic engineering by validating three neutral integration sites in Synechococcus sp. PCC 7002 |
| title_full_unstemmed | Streamlining recombination-mediated genetic engineering by validating three neutral integration sites in Synechococcus sp. PCC 7002 |
| title_short | Streamlining recombination-mediated genetic engineering by validating three neutral integration sites in Synechococcus sp. PCC 7002 |
| title_sort | streamlining recombination mediated genetic engineering by validating three neutral integration sites in synechococcus sp pcc 7002 |
| topic | Synechococcus sp. PCC7002 Neutral integration sites Genetic engineering Transformation BioBrick Cyanobacteria |
| url | http://link.springer.com/article/10.1186/s13036-017-0061-8 |
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