Cashra (cas9-facilitated homologous recombination assembly) method of constructing megabase-sized dna
文献类型：期刊论文
| 作者 | Zhou, Jianting1,2; Wu, Ronghai1,2; Xue, Xiaoli1; Qin, Zhongjun1 |
| 刊名 | Nucleic acids research
 |
| 出版日期 | 2016-08-19 |
| 卷号 | 44期号:14页码:9 |
| ISSN号 | 0305-1048 |
| DOI | 10.1093/nar/gkw475 |
| 通讯作者 | Xue, xiaoli(xlxue@sibs.ac.cn) ; Qin, zhongjun(qin@sibs.ac.cn) |
| 英文摘要 | Current dna assembly methods for preparing highly purified linear subassemblies require complex and time-consuming in vitro manipulations that hinder their ability to construct megabase-sized dnas (e.g. synthetic genomes). we have developed a new method designated 'cashra (cas9-facilitated homologous recombination assembly)' that directly uses large circular dnas in a one-step in vivo assembly process. the large circular dnas are co-introduced into saccharomyces cerevisiae by protoplast fusion, and they are cleaved by rna-guided cas9 nuclease to release the linear dna segments for subsequent assembly by the endogenous homologous recombination system. the cashra method allows efficient assembly of multiple large dna segments in vivo; thus, this approach should be useful in the last stage of genome construction. as a proof of concept, we combined cashra with an upstream assembly method (gibson procedure of genome assembly) and successfully constructed a 1.03 mb mge-syn1.0 (minimal genome of escherichia coli) that contained 449 essential genes and 267 important growth genes. we expect that cashra will be widely used in megabase-sized genome constructions. |
| WOS关键词 | TRANSFORMATION-ASSOCIATED RECOMBINATION ; SACCHAROMYCES-CEREVISIAE ; ESCHERICHIA-COLI ; ONE-STEP ; VIBRIO-CHOLERAE ; REDUCED-GENOME ; YEAST ; CRISPR ; GENE ; SYSTEMS |
| WOS研究方向 | Biochemistry & Molecular Biology |
| WOS类目 | Biochemistry & Molecular Biology |
| 语种 | 英语 |
| WOS记录号 | WOS:000382999900004 |
| 出版者 | OXFORD UNIV PRESS |
| URI标识 | http://www.irgrid.ac.cn/handle/1471x/2375638 |
| 专题 | 中国科学院大学 |
| 通讯作者 | Xue, Xiaoli; Qin, Zhongjun |
| 作者单位 | 1.Chinese Acad Sci, Shanghai Inst Biol Sci, Inst Plant Physiol & Ecol, Key Lab Synthet Biol, Shanghai 200032, Peoples R China 2.Univ Chinese Acad Sci, Beijing 100049, Peoples R China |
| 推荐引用方式 GB/T 7714 | Zhou, Jianting,Wu, Ronghai,Xue, Xiaoli,et al. Cashra (cas9-facilitated homologous recombination assembly) method of constructing megabase-sized dna[J]. Nucleic acids research,2016,44(14):9. |
| APA | Zhou, Jianting,Wu, Ronghai,Xue, Xiaoli,&Qin, Zhongjun.(2016).Cashra (cas9-facilitated homologous recombination assembly) method of constructing megabase-sized dna.Nucleic acids research,44(14),9. |
| MLA | Zhou, Jianting,et al."Cashra (cas9-facilitated homologous recombination assembly) method of constructing megabase-sized dna".Nucleic acids research 44.14(2016):9. |
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来源：中国科学院大学
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