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Reliable handling of highly A/T-rich genomic DNA for efficient generation of knockin strains of Dictyostelium discoideum
https://tokushima-u.repo.nii.ac.jp/records/2007334
https://tokushima-u.repo.nii.ac.jp/records/2007334b8a35653-e297-4475-941e-36e96e986d1b
| 名前 / ファイル | ライセンス | アクション |
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bmcbiotech_16_37.pdf (1.7 MB)
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| Item type | 文献 / Documents(1) | |||||||||||||||||||||||
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| 公開日 | 2020-03-26 | |||||||||||||||||||||||
| アクセス権 | ||||||||||||||||||||||||
| アクセス権 | open access | |||||||||||||||||||||||
| 資源タイプ | ||||||||||||||||||||||||
| 資源タイプ識別子 | http://purl.org/coar/resource_type/c_6501 | |||||||||||||||||||||||
| 資源タイプ | journal article | |||||||||||||||||||||||
| 出版社版DOI | ||||||||||||||||||||||||
| 関連識別子 | https://doi.org/10.1186/s12896-016-0267-8 | |||||||||||||||||||||||
| 関連名称 | 10.1186/s12896-016-0267-8 | |||||||||||||||||||||||
| 出版タイプ | ||||||||||||||||||||||||
| 出版タイプ | VoR | |||||||||||||||||||||||
| 出版タイプResource | http://purl.org/coar/version/c_970fb48d4fbd8a85 | |||||||||||||||||||||||
| タイトル | ||||||||||||||||||||||||
| タイトル | Reliable handling of highly A/T-rich genomic DNA for efficient generation of knockin strains of Dictyostelium discoideum | |||||||||||||||||||||||
| 著者 |
向井, あすか
× 向井, あすか
× イチラク, アヤ
× 堀川, 一樹
WEKO
586
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| 内容記述 | Background: Social amoeba, Dictyostelium discoideum, is a well-established model organism for studying cellular physiology and developmental pattern formation. Its haploid genome facilitates functional analysis of genes by a single round of mutagenesis including targeted disruption. Although the efficient generation of knockout strains based on an intrinsically high homologous recombination rate has been demonstrated, successful reports for knockin strains have been limited. As social amoeba has an exceptionally high adenine and thymine (A/T)-content, conventional plasmid-based vector construction has been constrained due to deleterious deletion in E. coli. Results: We describe here a simple and efficient strategy to construct GFP-knockin cassettes by using a linear DNA cloning vector derived from N15 bacteriophage. This allows reliable handling of DNA fragments whose A/T-content may be as high as 85 %, and which cannot be cloned into a circular plasmid. By optimizing the length of recombination arms, we successfully generate GFP-knockin strains for five genes involved in cAMP signalling, including a triple-colour knockin strain. Conclusions: This robust strategy would be useful in handling DNA fragments with biased A/T-contents such as the genome of lower organisms and the promoter/terminator regions of higher organisms. |
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| キーワード | ||||||||||||||||||||||||
| 主題 | A/T-rich genome | |||||||||||||||||||||||
| キーワード | ||||||||||||||||||||||||
| 主題 | Linear DNA cloning | |||||||||||||||||||||||
| キーワード | ||||||||||||||||||||||||
| 主題 | Knockin | |||||||||||||||||||||||
| キーワード | ||||||||||||||||||||||||
| 主題 | Dictyostelium discoideum | |||||||||||||||||||||||
| 書誌情報 |
en : BMC Biotechnology 巻 16, p. 37, 発行日 2016-04-14 |
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| 収録物ID | ||||||||||||||||||||||||
| 収録物識別子タイプ | ISSN | |||||||||||||||||||||||
| 収録物識別子 | 14726750 | |||||||||||||||||||||||
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| 収録物識別子タイプ | NCID | |||||||||||||||||||||||
| 収録物識別子 | AA12034730 | |||||||||||||||||||||||
| 出版者 | ||||||||||||||||||||||||
| 出版者 | BioMed Central | |||||||||||||||||||||||
| 出版者 | ||||||||||||||||||||||||
| 出版者 | Springer Nature | |||||||||||||||||||||||
| 権利情報 | ||||||||||||||||||||||||
| 権利情報 | © 2016 Mukai et al. Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. | |||||||||||||||||||||||
| EID | ||||||||||||||||||||||||
| 識別子 | 333986 | |||||||||||||||||||||||
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| 言語 | eng | |||||||||||||||||||||||
