"Perfect" designer chromosome V and behavior of a ring derivative.

Perfect matching of an assembled physical sequence to a specified designed sequence is crucial to verify design principles in genome synthesis. We designed and de novo synthesized 536,024-base pair chromosome synV in the "Build-A-Genome China" course. We corrected an initial isolate of synV to perfectly match the designed sequence using integrative cotransformation and clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9)-mediated editing in 22 steps; synV strains exhibit high fitness under a variety of culture conditions, compared with that of wild-type V strains.

Multigene Pathway Engineering with Regulatory Linkers (M-PERL).

Multigene pathway engineering usually needs amounts of part libraries on transcriptional and translational regulation as well as mutant enzymes to achieve the optimal part combinations of the target pathways. We report a new strategy for multigene pathway engineering with regulatory linkers (M-PERL) focusing on tuning the transcriptional start site (TSS) of yeast promoters. The regulatory linkers are composed of two homologous ends of two adjacent gene parts for assembly and a central regulatory region between them.

Engineering Yarrowia lipolytica for Campesterol Overproduction.

Campesterol is an important precursor for many sterol drugs, e.g. progesterone and hydrocortisone.