‘Gel-Stacks’ gently confine or reversibly immobilize arrays of single DNA molecules for manipulation and study
Large DNA molecules (>20 kb) are difficult analytes prone to breakage during serial manipulations and cannot be ‘rescued’ as full-length amplicons. Accordingly, to present, modify and analyze arrays of large, single DNA molecules, we created an easily realizable approach offering gentle confineme...
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| Format: | Article |
| Language: | English |
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Taylor & Francis Group
2024-06-01
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| Series: | BioTechniques |
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| Online Access: | https://www.tandfonline.com/doi/10.2144/btn-2023-0123 |
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| author | Susana Calle-Casteñeda Eamon Winden Alejandro Vasquez-Echeverri Matthew Schickling Evelyn Browning Juan Pablo Hernandez Ortiz David C Schwartz |
| author_facet | Susana Calle-Casteñeda Eamon Winden Alejandro Vasquez-Echeverri Matthew Schickling Evelyn Browning Juan Pablo Hernandez Ortiz David C Schwartz |
| author_sort | Susana Calle-Casteñeda |
| collection | DOAJ |
| description | Large DNA molecules (>20 kb) are difficult analytes prone to breakage during serial manipulations and cannot be ‘rescued’ as full-length amplicons. Accordingly, to present, modify and analyze arrays of large, single DNA molecules, we created an easily realizable approach offering gentle confinement conditions or immobilization via spermidine condensation for controlled delivery of reagents that support live imaging by epifluorescence microscopy termed ‘Gel-Stacks.’ Molecules are locally confined between two hydrogel surfaces without covalent tethering to support time-lapse imaging and multistep workflows that accommodate large DNA molecules. With a thin polyacrylamide gel layer covalently bound to a glass surface as the base and swappable, reagent-infused, agarose slabs on top, DNA molecules are stably presented for imaging during reagent delivery by passive diffusion. |
| format | Article |
| id | doaj-art-c3217d6ed5664cf7877aca2ea285586b |
| institution | Kabale University |
| issn | 0736-6205 1940-9818 |
| language | English |
| publishDate | 2024-06-01 |
| publisher | Taylor & Francis Group |
| record_format | Article |
| series | BioTechniques |
| spelling | doaj-art-c3217d6ed5664cf7877aca2ea285586b2025-01-16T15:38:47ZengTaylor & Francis GroupBioTechniques0736-62051940-98182024-06-0176628528910.2144/btn-2023-0123‘Gel-Stacks’ gently confine or reversibly immobilize arrays of single DNA molecules for manipulation and studySusana Calle-Casteñeda0Eamon Winden1Alejandro Vasquez-Echeverri2Matthew Schickling3Evelyn Browning4Juan Pablo Hernandez Ortiz5David C Schwartz6Laboratory for Molecular & Computational Genomics, University of Wisconsin–Madison, Madison, WI 53706, USALaboratory for Molecular & Computational Genomics, University of Wisconsin–Madison, Madison, WI 53706, USALaboratory for Molecular & Computational Genomics, University of Wisconsin–Madison, Madison, WI 53706, USALaboratory for Molecular & Computational Genomics, University of Wisconsin–Madison, Madison, WI 53706, USALaboratory for Molecular & Computational Genomics, University of Wisconsin–Madison, Madison, WI 53706, USAGHI One Health Colombia & One Health Genomic Laboratory, Universidad Nacional de Colombia – Medellín, Medellín, 050034, ColombiaLaboratory for Molecular & Computational Genomics, University of Wisconsin–Madison, Madison, WI 53706, USALarge DNA molecules (>20 kb) are difficult analytes prone to breakage during serial manipulations and cannot be ‘rescued’ as full-length amplicons. Accordingly, to present, modify and analyze arrays of large, single DNA molecules, we created an easily realizable approach offering gentle confinement conditions or immobilization via spermidine condensation for controlled delivery of reagents that support live imaging by epifluorescence microscopy termed ‘Gel-Stacks.’ Molecules are locally confined between two hydrogel surfaces without covalent tethering to support time-lapse imaging and multistep workflows that accommodate large DNA molecules. With a thin polyacrylamide gel layer covalently bound to a glass surface as the base and swappable, reagent-infused, agarose slabs on top, DNA molecules are stably presented for imaging during reagent delivery by passive diffusion.https://www.tandfonline.com/doi/10.2144/btn-2023-0123confinementepifluorescence microscopyFick's second lawhydrogelslarge DNA molecule arraysreagent transport |
| spellingShingle | Susana Calle-Casteñeda Eamon Winden Alejandro Vasquez-Echeverri Matthew Schickling Evelyn Browning Juan Pablo Hernandez Ortiz David C Schwartz ‘Gel-Stacks’ gently confine or reversibly immobilize arrays of single DNA molecules for manipulation and study BioTechniques confinement epifluorescence microscopy Fick's second law hydrogels large DNA molecule arrays reagent transport |
| title | ‘Gel-Stacks’ gently confine or reversibly immobilize arrays of single DNA molecules for manipulation and study |
| title_full | ‘Gel-Stacks’ gently confine or reversibly immobilize arrays of single DNA molecules for manipulation and study |
| title_fullStr | ‘Gel-Stacks’ gently confine or reversibly immobilize arrays of single DNA molecules for manipulation and study |
| title_full_unstemmed | ‘Gel-Stacks’ gently confine or reversibly immobilize arrays of single DNA molecules for manipulation and study |
| title_short | ‘Gel-Stacks’ gently confine or reversibly immobilize arrays of single DNA molecules for manipulation and study |
| title_sort | gel stacks gently confine or reversibly immobilize arrays of single dna molecules for manipulation and study |
| topic | confinement epifluorescence microscopy Fick's second law hydrogels large DNA molecule arrays reagent transport |
| url | https://www.tandfonline.com/doi/10.2144/btn-2023-0123 |
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