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Science. 2017 Mar 10;355(6329). pii: eaaf4706. doi: 10.1126/science.aaf4706.

Bug mapping and fitness testing of chemically synthesized chromosome X.

Wu Y1,2, Li BZ1,2, Zhao M1,2, Mitchell LA3, Xie ZX1,2, Lin QH1,2, Wang X1,2, Xiao WH1,2, Wang Y1,2, Zhou X1,2, Liu H1,2, Li X1,2, Ding MZ1,2, Liu D1,2, Zhang L1,2, Liu BL1,2, Wu XL1,2, Li FF1,2, Dong XT1,2, Jia B1,2, Zhang WZ1,2, Jiang GZ1,2, Liu Y1,2, Bai X1,2, Song TQ1,2, Chen Y1,2, Zhou SJ1,2, Zhu RY1,2, Gao F1,2, Kuang Z3, Wang X3, Shen M3, Yang K4, Stracquadanio G4,5, Richardson SM4, Lin Y6, Wang L6, Walker R7, Luo Y7, Ma PS1,2, Yang H8,9, Cai Y7, Dai J6, Bader JS4, Boeke JD3, Yuan YJ10,2.

Author information

1
Key Laboratory of Systems Bioengineering (Ministry of Education), School of Chemical Engineering and Technology, Tianjin University, Tianjin, 300072, PR China.
2
SynBio Research Platform, Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), Tianjin University, Tianjin, 300072, PR China.
3
Institute for Systems Genetics and Department of Biochemistry and Molecular Pharmacology, New York University (NYU) Langone Medical Center, New York City, NY 10016, USA.
4
High Throughput Biology Center and Department of Biomedical Engineering, Johns Hopkins University, Baltimore, MD 21205, USA.
5
School of Computer Science and Electronic Engineering, University of Essex, Wivenhoe Park, Colchester CO4 3SQ, UK.
6
Key laboratory of Industrial Biocatalysis (Ministry of Education), Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing, 100084, PR China.
7
School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3BF, Scotland, UK.
8
BGI-Shenzhen, Shenzhen, 518083, PR China.
9
James D. Watson Institute of Genome Sciences, Hangzhou 310058, PR China.
10
Key Laboratory of Systems Bioengineering (Ministry of Education), School of Chemical Engineering and Technology, Tianjin University, Tianjin, 300072, PR China. yjyuan@tju.edu.cn.

Abstract

Debugging a genome sequence is imperative for successfully building a synthetic genome. As part of the effort to build a designer eukaryotic genome, yeast synthetic chromosome X (synX), designed as 707,459 base pairs, was synthesized chemically. SynX exhibited good fitness under a wide variety of conditions. A highly efficient mapping strategy called pooled PCRTag mapping (PoPM), which can be generalized to any watermarked synthetic chromosome, was developed to identify genetic alterations that affect cell fitness ("bugs"). A series of bugs were corrected that included a large region bearing complex amplifications, a growth defect mapping to a recoded sequence in FIP1, and a loxPsym site affecting promoter function of ATP2 PoPM is a powerful tool for synthetic yeast genome debugging and an efficient strategy for phenotype-genotype mapping.

PMID:
28280152
PMCID:
PMC5679077
DOI:
10.1126/science.aaf4706
[Indexed for MEDLINE]
Free PMC Article

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