A simple negative selection method to identify adenovirus recombinants using colony PCR
Background: The AdEasy system is a fast-track system for generating recombinant adenoviruses using the efficient homologous recombination machinery between shuttle and adenovirus backbone plasmids in Escherichia coli BJ5183 cells. The key step is homologous recombination in BJ5183 cells, which is dr...
| Main Authors: | , , , |
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| Format: | Article |
| Language: | English |
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Elsevier
2014-01-01
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| Series: | Electronic Journal of Biotechnology |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S0717345813000092 |
| _version_ | 1818890902620340224 |
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| author | Yongliang Zhao Zongming Dong Tengfei Li Gang Huang |
| author_facet | Yongliang Zhao Zongming Dong Tengfei Li Gang Huang |
| author_sort | Yongliang Zhao |
| collection | DOAJ |
| description | Background: The AdEasy system is a fast-track system for generating recombinant adenoviruses using the efficient homologous recombination machinery between shuttle and adenovirus backbone plasmids in Escherichia coli BJ5183 cells. The key step is homologous recombination in BJ5183 cells, which is driven by RecA activity. However, culture time is stringently limited to reduce the damage to recombinant plasmids by RecA activity. Therefore, rapid identification of recombinant adenoviruses within the limited time-period is critical.
Results: We developed a simple negative selection method to identify recombinant adenoviruses using colony PCR, which improves the efficiency of adenovirus recombination screening and packaging.
Conclusions: The negative selection method to identify AdEasy adenovirus recombinants by colony PCR can identify the recombined colony within a short time-period, and maximally avoid damage to the recombinant plasmid by limiting recombination time, resulting in improved adenovirus packaging. |
| first_indexed | 2024-12-19T17:32:18Z |
| format | Article |
| id | doaj.art-3dd5636324d74de89c580ce6b027cf79 |
| institution | Directory Open Access Journal |
| issn | 0717-3458 |
| language | English |
| last_indexed | 2024-12-19T17:32:18Z |
| publishDate | 2014-01-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Electronic Journal of Biotechnology |
| spelling | doaj.art-3dd5636324d74de89c580ce6b027cf792022-12-21T20:12:25ZengElsevierElectronic Journal of Biotechnology0717-34582014-01-01171464910.1016/j.ejbt.2013.12.008A simple negative selection method to identify adenovirus recombinants using colony PCRYongliang Zhao0Zongming Dong1Tengfei Li2Gang Huang3Department of General Surgery, Southwest Hospital, Third Military Medical University, Chongqing 400038, ChinaDepartment of General Surgery, Southwest Hospital, Third Military Medical University, Chongqing 400038, ChinaDepartment of General Surgery, Southwest Hospital, Third Military Medical University, Chongqing 400038, ChinaDepartment of Medical Genetics, College of Basic Medicine, Third Military Medical University, Chongqing 400038, ChinaBackground: The AdEasy system is a fast-track system for generating recombinant adenoviruses using the efficient homologous recombination machinery between shuttle and adenovirus backbone plasmids in Escherichia coli BJ5183 cells. The key step is homologous recombination in BJ5183 cells, which is driven by RecA activity. However, culture time is stringently limited to reduce the damage to recombinant plasmids by RecA activity. Therefore, rapid identification of recombinant adenoviruses within the limited time-period is critical. Results: We developed a simple negative selection method to identify recombinant adenoviruses using colony PCR, which improves the efficiency of adenovirus recombination screening and packaging. Conclusions: The negative selection method to identify AdEasy adenovirus recombinants by colony PCR can identify the recombined colony within a short time-period, and maximally avoid damage to the recombinant plasmid by limiting recombination time, resulting in improved adenovirus packaging.http://www.sciencedirect.com/science/article/pii/S0717345813000092AdenovirusAdEasyColony PCRHomologous recombinationRecA |
| spellingShingle | Yongliang Zhao Zongming Dong Tengfei Li Gang Huang A simple negative selection method to identify adenovirus recombinants using colony PCR Electronic Journal of Biotechnology Adenovirus AdEasy Colony PCR Homologous recombination RecA |
| title | A simple negative selection method to identify adenovirus recombinants using colony PCR |
| title_full | A simple negative selection method to identify adenovirus recombinants using colony PCR |
| title_fullStr | A simple negative selection method to identify adenovirus recombinants using colony PCR |
| title_full_unstemmed | A simple negative selection method to identify adenovirus recombinants using colony PCR |
| title_short | A simple negative selection method to identify adenovirus recombinants using colony PCR |
| title_sort | simple negative selection method to identify adenovirus recombinants using colony pcr |
| topic | Adenovirus AdEasy Colony PCR Homologous recombination RecA |
| url | http://www.sciencedirect.com/science/article/pii/S0717345813000092 |
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