Streamlining standard bacteriophage methods for higher throughput
A universal tool in the culture-based study of bacterial viruses (bacteriophages, or phages) is the agar overlay, which is used in the isolation of new viruses, and in their quantification and purification. Here, simple optimizations that increase efficiency and throughput in agar overlay based isol...
| Main Authors: | , |
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| Format: | Article |
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Elsevier BV
2018
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| Online Access: | http://hdl.handle.net/1721.1/117419 https://orcid.org/0000-0002-1997-947X https://orcid.org/0000-0001-9296-3733 |
| _version_ | 1811091129316147200 |
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| author | Kauffman, Anne Kathryn Marie Polz, Martin F |
| author2 | Massachusetts Institute of Technology. Center for Microbiome Informatics and Therapeutics |
| author_facet | Massachusetts Institute of Technology. Center for Microbiome Informatics and Therapeutics Kauffman, Anne Kathryn Marie Polz, Martin F |
| author_sort | Kauffman, Anne Kathryn Marie |
| collection | MIT |
| description | A universal tool in the culture-based study of bacterial viruses (bacteriophages, or phages) is the agar overlay, which is used in the isolation of new viruses, and in their quantification and purification. Here, simple optimizations that increase efficiency and throughput in agar overlay based isolation and cultivation of virus-host systems are presented. The agar overlay is streamlined to minimize steps and materials. Serial purification of viruses from viral colonies (plaques) is optimized to eliminate steps by combining purification by serial re-streaking with the optimized agar overlay approach. Finally, recommendations are made for efficient archival and storage of virus plaques. In sum, this work presents: • Tube-free Agar Overlays: rapid plaque assays with fewer steps and materials• Molten Streaking for Singles: rapid tube-free serial purification of viruses• Archiving Plaques: saving virus purification for later Keywords: Agar overlay; Agar layer; Top agar; Bottom agar; Plaque; Virus; Phage; Isolation; Cultivation; Purification |
| first_indexed | 2024-09-23T14:57:28Z |
| format | Article |
| id | mit-1721.1/117419 |
| institution | Massachusetts Institute of Technology |
| last_indexed | 2024-09-23T14:57:28Z |
| publishDate | 2018 |
| publisher | Elsevier BV |
| record_format | dspace |
| spelling | mit-1721.1/1174192022-10-01T23:35:06Z Streamlining standard bacteriophage methods for higher throughput Kauffman, Anne Kathryn Marie Polz, Martin F Massachusetts Institute of Technology. Center for Microbiome Informatics and Therapeutics Massachusetts Institute of Technology. Department of Civil and Environmental Engineering Kauffman, Anne Kathryn Marie Polz, Martin F A universal tool in the culture-based study of bacterial viruses (bacteriophages, or phages) is the agar overlay, which is used in the isolation of new viruses, and in their quantification and purification. Here, simple optimizations that increase efficiency and throughput in agar overlay based isolation and cultivation of virus-host systems are presented. The agar overlay is streamlined to minimize steps and materials. Serial purification of viruses from viral colonies (plaques) is optimized to eliminate steps by combining purification by serial re-streaking with the optimized agar overlay approach. Finally, recommendations are made for efficient archival and storage of virus plaques. In sum, this work presents: • Tube-free Agar Overlays: rapid plaque assays with fewer steps and materials• Molten Streaking for Singles: rapid tube-free serial purification of viruses• Archiving Plaques: saving virus purification for later Keywords: Agar overlay; Agar layer; Top agar; Bottom agar; Plaque; Virus; Phage; Isolation; Cultivation; Purification National Science Foundation (U.S.) (Grant OCE-1435993) 2018-08-20T17:30:02Z 2018-08-20T17:30:02Z 2018-01 2017-10 2018-08-20T14:35:10Z Article http://purl.org/eprint/type/JournalArticle 2215-0161 http://hdl.handle.net/1721.1/117419 Kauffman, Kathryn M. and Martin F. Polz. “Streamlining Standard Bacteriophage Methods for Higher Throughput.” MethodsX 5 (2018): 159–172 © 2018 The Author(s) https://orcid.org/0000-0002-1997-947X https://orcid.org/0000-0001-9296-3733 http://dx.doi.org/10.1016/J.MEX.2018.01.007 MethodsX Creative Commons Attribution 4.0 International License http://creativecommons.org/licenses/by/4.0/ application/pdf Elsevier BV Elsevier |
| spellingShingle | Kauffman, Anne Kathryn Marie Polz, Martin F Streamlining standard bacteriophage methods for higher throughput |
| title | Streamlining standard bacteriophage methods for higher throughput |
| title_full | Streamlining standard bacteriophage methods for higher throughput |
| title_fullStr | Streamlining standard bacteriophage methods for higher throughput |
| title_full_unstemmed | Streamlining standard bacteriophage methods for higher throughput |
| title_short | Streamlining standard bacteriophage methods for higher throughput |
| title_sort | streamlining standard bacteriophage methods for higher throughput |
| url | http://hdl.handle.net/1721.1/117419 https://orcid.org/0000-0002-1997-947X https://orcid.org/0000-0001-9296-3733 |
| work_keys_str_mv | AT kauffmanannekathrynmarie streamliningstandardbacteriophagemethodsforhigherthroughput AT polzmartinf streamliningstandardbacteriophagemethodsforhigherthroughput |