期刊
ACS SYNTHETIC BIOLOGY
卷 4, 期 3, 页码 213-220出版社
AMER CHEMICAL SOC
DOI: 10.1021/sb500241e
关键词
in vivo assembly; synthetic yeast genome; Sc2.0; Build-A-Genome; synthetic biology
资金
- Chinese National High Technology Research and Development Program (863 Program) [2012AA02A701]
- National Basic Research Program of China (973 Program) [2014CB745100]
- Johns Hopkins Yeung Center
- US National Science Foundation SAVI program [MCB-1158201]
- PDF from NSERC
- Direct For Biological Sciences
- Div Of Molecular and Cellular Bioscience [1443299] Funding Source: National Science Foundation
- Direct For Biological Sciences
- Div Of Molecular and Cellular Bioscience [1445545, 1441866] Funding Source: National Science Foundation
We describe rapid assembly of DNA overlapping multifragments (RADOM), an improved assembly method via homologous recombination in Saccharomyces cerevisiae, which combines assembly in yeas to with blue/white screening in Escherichia coli. We show that RADOM can successfully assemble similar to 3 and similar to 10 kb DNA fragments that are highly similar to the yeast genome rapidly and accurately. This method was tested in the Build-A-Genome course by undergraduate students, where 125 similar to 3 kb minichunks from the synthetic yeast genome project Sc2.0 were assembled. Here, 122 out of 125 minichunks achieved insertions with correct sizes, and 102 minichunks were sequenced verified. As this method reduces the time-consuming and labor-intensive efforts of yeast assembly by improving the screening efficiency for correct assemblies, it may find routine applications in the construction of DNA fragments, especially in hierarchical assembly projects.
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