Molecular Genetics of Actinomycetes and Synthetic Microbiology
My laboratoty has worked on biological functions (e.g., replication, transfer and evolution etc.) of Streptomyces linear and circular plasmids and their applications for many years. We have extended the interest to develop genetic tools and Streptomyces genomics, especially genomic engineering. We also constructed and genomic modified the antibiotics industrial strains (e.g., doramectin) to improve their properties via collaborated with pharmaceutical companies (e.g., Zhejiang Hisun). We initiated a big project on the minimal genome in year 2011. Currently, we have made progresses on reconstruction of the simplified Escherichia coli genomes by using both bottom-up and top-down strategies, development of genomic tools, and artificially clustering and modulating of some essential functional genes, etc.
Principal Investigator: Dr. Zhong-Jun Qin, Professor. Email: firstname.lastname@example.org
Staff: Li Zhong, XiaoLi Xue, HaiYang Xia, Tao Wang, Peng Jiang
Postdoctoral fellow: Jianting Zhou
Graduate Students: YangYang Shao, Min Zhou, RongHai Wu, Ting He, Xiaoshu Ma, Guanjun Qiao, Ning Lu
Recent research progress
Construction of an E.coli chassis cell with simplified genome
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 cointroduced 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 ofEscherichia coli) that contained 449 essential genes and 267 important growth genes. We expect that CasHRA will be widely used in megabase-sized genome constructions.
- Jianting Zhou, Ronghai Wu, Xiaoli Xue and Zhongjun Qin*. (2016) CasHRA (Cas9-facilitated Homologous Recombination Assembly) method of constructing megabase-sized DNA. Nucleic Acids Research. 44(14):e124.
- Chen J, Xia HY*, Dang FJ, Xu QY, Li WJ, Qin ZJ* (2015) Characterization of the chromosomal integration of Saccharopolyspora plasmid pCM32 and its application to improve production of spinosyn in Saccharopolysporaspinosa. ApplMicrobiol Biotech. 99:10141–10149.
- Xue XL*, Wang T, Jiang P, Shao YY, Zhou M, Zhong L, Wu RH, Zhou JT, Xia HY, Zhao GP, Qin ZJ* (2015) MEGA (Multiple Essential Genes Assembling) deletion and replacement method for genome reduction in Escherichia coli. ACS Syn Biol. 4:700–706.