The study of protein recruitment to laser-induced DNA lesions can be distorted by photoconversion of the DNA binding dye Hoechst [version 2; peer review: 1 approved, 2 approved with reservations]
A commonly used approach for assessing DNA repair factor recruitment in mammalian cells is to induce DNA damage with a laser in the UV or near UV range and follow the local increase of GFP-tagged proteins at the site of damage. Often these measurements are performed in the presence of the blue DNA d...
| Main Authors: | , |
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| Format: | Article |
| Language: | English |
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F1000 Research Ltd
2019-04-01
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| Series: | F1000Research |
| Online Access: | https://f1000research.com/articles/8-104/v2 |
| _version_ | 1818943311659925504 |
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| author | Verena Hurst Susan M. Gasser |
| author_facet | Verena Hurst Susan M. Gasser |
| author_sort | Verena Hurst |
| collection | DOAJ |
| description | A commonly used approach for assessing DNA repair factor recruitment in mammalian cells is to induce DNA damage with a laser in the UV or near UV range and follow the local increase of GFP-tagged proteins at the site of damage. Often these measurements are performed in the presence of the blue DNA dye Hoechst, which is used as a photosensitizer. However, a light-induced switch of Hoechst from a blue-light to a green-light emitter will give a false positive signal at the site of damage. Thus, photoconversion signals must be subtracted from the overall green-light emission to determine true recruitment. Here we demonstrate the photoconversion effect and suggest control experiments to exclude false-positive results. |
| first_indexed | 2024-12-20T07:25:19Z |
| format | Article |
| id | doaj.art-56dbe9deae70473ca6abceab4088a0b4 |
| institution | Directory Open Access Journal |
| issn | 2046-1402 |
| language | English |
| last_indexed | 2024-12-20T07:25:19Z |
| publishDate | 2019-04-01 |
| publisher | F1000 Research Ltd |
| record_format | Article |
| series | F1000Research |
| spelling | doaj.art-56dbe9deae70473ca6abceab4088a0b42022-12-21T19:48:33ZengF1000 Research LtdF1000Research2046-14022019-04-01810.12688/f1000research.17865.220618The study of protein recruitment to laser-induced DNA lesions can be distorted by photoconversion of the DNA binding dye Hoechst [version 2; peer review: 1 approved, 2 approved with reservations]Verena Hurst0Susan M. Gasser1Friedrich Miescher Institute for Biomedical Research, Maulbeerstrasse 66, Basel, CH-4058, SwitzerlandFriedrich Miescher Institute for Biomedical Research, Maulbeerstrasse 66, Basel, CH-4058, SwitzerlandA commonly used approach for assessing DNA repair factor recruitment in mammalian cells is to induce DNA damage with a laser in the UV or near UV range and follow the local increase of GFP-tagged proteins at the site of damage. Often these measurements are performed in the presence of the blue DNA dye Hoechst, which is used as a photosensitizer. However, a light-induced switch of Hoechst from a blue-light to a green-light emitter will give a false positive signal at the site of damage. Thus, photoconversion signals must be subtracted from the overall green-light emission to determine true recruitment. Here we demonstrate the photoconversion effect and suggest control experiments to exclude false-positive results.https://f1000research.com/articles/8-104/v2 |
| spellingShingle | Verena Hurst Susan M. Gasser The study of protein recruitment to laser-induced DNA lesions can be distorted by photoconversion of the DNA binding dye Hoechst [version 2; peer review: 1 approved, 2 approved with reservations] F1000Research |
| title | The study of protein recruitment to laser-induced DNA lesions can be distorted by photoconversion of the DNA binding dye Hoechst [version 2; peer review: 1 approved, 2 approved with reservations] |
| title_full | The study of protein recruitment to laser-induced DNA lesions can be distorted by photoconversion of the DNA binding dye Hoechst [version 2; peer review: 1 approved, 2 approved with reservations] |
| title_fullStr | The study of protein recruitment to laser-induced DNA lesions can be distorted by photoconversion of the DNA binding dye Hoechst [version 2; peer review: 1 approved, 2 approved with reservations] |
| title_full_unstemmed | The study of protein recruitment to laser-induced DNA lesions can be distorted by photoconversion of the DNA binding dye Hoechst [version 2; peer review: 1 approved, 2 approved with reservations] |
| title_short | The study of protein recruitment to laser-induced DNA lesions can be distorted by photoconversion of the DNA binding dye Hoechst [version 2; peer review: 1 approved, 2 approved with reservations] |
| title_sort | study of protein recruitment to laser induced dna lesions can be distorted by photoconversion of the dna binding dye hoechst version 2 peer review 1 approved 2 approved with reservations |
| url | https://f1000research.com/articles/8-104/v2 |
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