Li Zhen-Hai, Liu Min, Wang Feng-Qing, Wei Dong-Zhi
State Key Laboratory of Bioreactor Engineering, Newworld Institute of Biotechnology, East China University of Science and Technology, Shanghai, 200237, People's Republic of China.
Biotechnol Lett. 2018 Aug;40(8):1253-1261. doi: 10.1007/s10529-018-2574-8. Epub 2018 May 24.
To test the applicability of Cpf1 from Francisella novivida in genomic integration of in vivo assembled DNA parts in Saccharomyces cerevisiae.
An easy-to-use vector toolkit, containing a CEN6/ARS4 plasmid expressing Cpf1 from Francisella novivida (FnCpf1) and a 2 μ plasmid for crRNA or crRNA array expressing, was constructed for Cpf1-assisted genomic integration in S. cerevisiae. Our results showed that FnCpf1 allowed for targeted singleplex, doubleplex, and tripleplex genomic integration of in vivo assembled DNA parts with efficiencies of 95, 52, and 43%, respectively.
CRISPR-Cpf1 system allows for efficient genomic integration of in vivo assembled DNA parts in S. cerevisiae, and thus provides an alternative CRISPR-Cas method for metabolic pathway engineering in addition to CRISPR-Cas9 system previously reported for yeast.
测试来自新凶手弗朗西斯菌的Cpf1在酿酒酵母体内组装DNA片段的基因组整合中的适用性。
构建了一个易于使用的载体工具包，其中包含一个表达来自新凶手弗朗西斯菌Cpf1（FnCpf1）的CEN6/ARS4质粒和一个用于表达crRNA或crRNA阵列的2μ质粒，用于在酿酒酵母中进行Cpf1辅助的基因组整合。我们的结果表明，FnCpf1能够实现体内组装DNA片段的靶向单重、双重和三重基因组整合，效率分别为95%、52%和43%。
CRISPR-Cpf1系统能够在酿酒酵母中实现体内组装DNA片段的高效基因组整合，因此除了先前报道的用于酵母的CRISPR-Cas9系统外，还为代谢途径工程提供了另一种CRISPR-Cas方法。
