The TelN/tos-assisted precise targeting of chromosome segments (TAPE)
Introduction: Performing genomic large segmentation experiments will promote the annotation of complex genomic functions and contribute to the synthesis of designed genomes. It is challenging to obtain and manipulate large or complex DNA sequences with high efficiency. Objectives: This study aims to...
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| Language: | English |
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Elsevier
2022-11-01
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| Series: | Journal of Advanced Research |
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| Online Access: | http://www.sciencedirect.com/science/article/pii/S2090123222000303 |
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| author | You-Zhi Cui Jian-Ting Zhou Bing-Zhi Li Ying-Jin Yuan |
| author_facet | You-Zhi Cui Jian-Ting Zhou Bing-Zhi Li Ying-Jin Yuan |
| author_sort | You-Zhi Cui |
| collection | DOAJ |
| description | Introduction: Performing genomic large segmentation experiments will promote the annotation of complex genomic functions and contribute to the synthesis of designed genomes. It is challenging to obtain and manipulate large or complex DNA sequences with high efficiency. Objectives: This study aims to develop an effective method for direct cloning of target genome sequences from different species. Methods: The TelN/tos system and a linear plasmid vector were first used to directly clone the large genomic segments in E. coli. For the in vitro cloning reaction, two telomeric sites were developed using TelN protelomerase at the end of the linear plasmid vector. The target DNA sequence can be easily hooked with the homology arms and maintained as a linear artificial chromosome with arbitrary restriction sites in a specific E. coli strain. Results: Using the linear cloning strategy, we successfully cloned the bacterial DNA fragment of 156 kb, a yeast genomic fragment of 124 kb and mammalian mitochondrial fragment of 16 kb. The results showed a considerable improvement in cloning efficiency and demonstrated the important role of vector ratio in the cloning process. Conclusion: Due to the high efficiency and stability, TAPE is an effective technique for DNA cloning and fundamental molecular biotechnology method in synthetic biology. |
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| format | Article |
| id | doaj.art-b008fbcacb18433f9228284746e1d920 |
| institution | Directory Open Access Journal |
| issn | 2090-1232 |
| language | English |
| last_indexed | 2024-04-11T23:38:20Z |
| publishDate | 2022-11-01 |
| publisher | Elsevier |
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| series | Journal of Advanced Research |
| spelling | doaj.art-b008fbcacb18433f9228284746e1d9202022-12-22T03:56:53ZengElsevierJournal of Advanced Research2090-12322022-11-0141169177The TelN/tos-assisted precise targeting of chromosome segments (TAPE)You-Zhi Cui0Jian-Ting Zhou1Bing-Zhi Li2Ying-Jin Yuan3Frontiers Science Center for Synthetic Biology and Key Laboratory of Systems Bioengineering (Ministry of Education), School of Chemical Engineering and Technology, Tianjin University, 300072 Tianjin, China; Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), Tianjin University, 300072 Tianjin, ChinaFrontiers Science Center for Synthetic Biology and Key Laboratory of Systems Bioengineering (Ministry of Education), School of Chemical Engineering and Technology, Tianjin University, 300072 Tianjin, China; Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), Tianjin University, 300072 Tianjin, ChinaCorresponding author at: Frontiers Science Center for Synthetic Biology and Key Laboratory of Systems Bioengineering (Ministry of Education), School of Chemical Engineering and Technology, Tianjin University, 300072 Tianjin, China.; Frontiers Science Center for Synthetic Biology and Key Laboratory of Systems Bioengineering (Ministry of Education), School of Chemical Engineering and Technology, Tianjin University, 300072 Tianjin, China; Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), Tianjin University, 300072 Tianjin, ChinaFrontiers Science Center for Synthetic Biology and Key Laboratory of Systems Bioengineering (Ministry of Education), School of Chemical Engineering and Technology, Tianjin University, 300072 Tianjin, China; Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), Tianjin University, 300072 Tianjin, ChinaIntroduction: Performing genomic large segmentation experiments will promote the annotation of complex genomic functions and contribute to the synthesis of designed genomes. It is challenging to obtain and manipulate large or complex DNA sequences with high efficiency. Objectives: This study aims to develop an effective method for direct cloning of target genome sequences from different species. Methods: The TelN/tos system and a linear plasmid vector were first used to directly clone the large genomic segments in E. coli. For the in vitro cloning reaction, two telomeric sites were developed using TelN protelomerase at the end of the linear plasmid vector. The target DNA sequence can be easily hooked with the homology arms and maintained as a linear artificial chromosome with arbitrary restriction sites in a specific E. coli strain. Results: Using the linear cloning strategy, we successfully cloned the bacterial DNA fragment of 156 kb, a yeast genomic fragment of 124 kb and mammalian mitochondrial fragment of 16 kb. The results showed a considerable improvement in cloning efficiency and demonstrated the important role of vector ratio in the cloning process. Conclusion: Due to the high efficiency and stability, TAPE is an effective technique for DNA cloning and fundamental molecular biotechnology method in synthetic biology.http://www.sciencedirect.com/science/article/pii/S2090123222000303Large genomic segmentsTargeted cloningTelN/tos systemLinear artificial chromosomes |
| spellingShingle | You-Zhi Cui Jian-Ting Zhou Bing-Zhi Li Ying-Jin Yuan The TelN/tos-assisted precise targeting of chromosome segments (TAPE) Journal of Advanced Research Large genomic segments Targeted cloning TelN/tos system Linear artificial chromosomes |
| title | The TelN/tos-assisted precise targeting of chromosome segments (TAPE) |
| title_full | The TelN/tos-assisted precise targeting of chromosome segments (TAPE) |
| title_fullStr | The TelN/tos-assisted precise targeting of chromosome segments (TAPE) |
| title_full_unstemmed | The TelN/tos-assisted precise targeting of chromosome segments (TAPE) |
| title_short | The TelN/tos-assisted precise targeting of chromosome segments (TAPE) |
| title_sort | teln tos assisted precise targeting of chromosome segments tape |
| topic | Large genomic segments Targeted cloning TelN/tos system Linear artificial chromosomes |
| url | http://www.sciencedirect.com/science/article/pii/S2090123222000303 |
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