Pursuing construction of the synthetic eukaryotic chromosome perfectly matching the designer sequence, the research team overcame significant obstacles. With all the 3331bp randomly distributed mutations and other unpredictable events corrected in the designed sequence and performing with a similar fitness compared with native strains, the completed synV chromosome supported the underlying design principles of Sc2.0. After the paper was originally submitted, the team continued their corrections, leading to the title of the synV article being changed from "approaching a variant-free designer chromosome" to "'perfect' designer chromosome V". "Together with my colleagues, I spent almost 18 months mapping the bugs and correcting errors that led to the yeast being non-viable or having growth defects," says Xie, describing the study of yeast chromosome synV.

Aiming to provide a model with which to study human ring chromosome disorders, the linear synV was converted to a ring synthetic chromosome, and it can extend genome design principles, as well as providing an insight into genomic rearrangement and ring chromosome evolution.

Certain genetic alternations in the chromosome would affect cell fitness and cause "bugs"; debugging is therefore imperative for successfully building a synthetic genome. The malfunction of synthetic genomes remains one of the most common obstacles, as it is difficult to uncover the underlying causes. In a bid to tackle this problem, TJU developed a highly efficient debugging method called pooled PCRTag mapping (PoPM) by using a pooling strategy and the Sc2.0 PCRTag system, which is generalized to watermarked synthetic chromosomes. PoPM is a powerful tool for synthetic yeast genome debugging which accelerated the progress of the whole Sc2.0 project. "During synX assembly, the PoPM method was applied to identify and eliminate genetic alternations that affect cell fitness, sequences we refer to as 'bugs', and several details of yeast biology were uncovered by debugging. PoPM represents an efficient strategy for phenotype-genotype mapping which will be applicable for expanding understanding of yeast genomic and cellular functions," Wu said.

Characterization of synX and debugging by pooled PCRTag mapping. (Top) Design overview of synthetic chromosome X. (Bottom) Flow diagram of pooled PCRTag mapping (PoPM).

Nominated by the Ministry of Science and Technology of the People's Republic of China, Tianjin University became the National Base for Synthetic Biotechnology International Science and Technology Cooperation in 2013. The Synthetic Biology major, established at Tianjin University in 2014, has been sustainably fostering a young generation of synthetic biology researchers.

Tianjin University synthetic biology team has started a new Long March towards providing solutions to societal challenges, assembling super-large designer genomes to improve human health, prevent and cure diseases, provide clean energy, and promote a sustainable environment.