Bug mapping and fitness testing of chemically synthesized chromosome X

Yi Wu; Bing Zhi Li; Meng Zhao; Leslie A. Mitchell; Ze Xiong Xie; Qiu Hui Lin; Xia Wang; Wen Hai Xiao; Ying Wang; Xiao Zhou; Hong Liu; Xia Li; Ming Zhu Ding; Duo Liu; Lu Zhang; Bao Li Liu; Xiao Le Wu; Fei Fei Li; Xiu Tao Dong; Bin Jia; Wen Zheng Zhang; Guo Zhen Jiang; Yue Liu; Xue Bai; Tian Qing Song; Yan Chen; Si Jie Zhou; Rui Ying Zhu; Feng Gao; Zheng Kuang; Xuya Wang; Michael Shen; Kun Yang; Giovanni Stracquadanio; Sarah M. Richardson; Yicong Lin; Lihui Wang; Roy Walker; Yisha Luo; Ping Sheng Ma; Huanming Yang; Yizhi Cai; Junbiao Dai; Joel S. Bader; Jef D. Boeke; Ying Jin Yuan

doi:10.1126/science.aaf4706

Bug mapping and fitness testing of chemically synthesized chromosome X

Yi Wu, Bing Zhi Li, Meng Zhao, Leslie A. Mitchell, Ze Xiong Xie, Qiu Hui Lin, Xia Wang, Wen Hai Xiao, Ying Wang, Xiao Zhou, Hong Liu, Xia Li, Ming Zhu Ding, Duo Liu, Lu Zhang, Bao Li Liu, Xiao Le Wu, Fei Fei Li, Xiu Tao Dong, Bin JiaWen Zheng Zhang, Guo Zhen Jiang, Yue Liu, Xue Bai, Tian Qing Song, Yan Chen, Si Jie Zhou, Rui Ying Zhu, Feng Gao, Zheng Kuang, Xuya Wang, Michael Shen, Kun Yang, Giovanni Stracquadanio, Sarah M. Richardson, Yicong Lin, Lihui Wang, Roy Walker, Yisha Luo, Ping Sheng Ma, Huanming Yang, Yizhi Cai, Junbiao Dai, Joel S. Bader, Jef D. Boeke, Ying Jin Yuan

Biomedical Engineering

Research output: Contribution to journal › Article › peer-review

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.

Original language	English (US)
Article number	eaaf4706
Journal	Science
Volume	355
Issue number	6329
DOIs	https://doi.org/10.1126/science.aaf4706
State	Published - Mar 10 2017

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Wu, Y., Li, B. Z., Zhao, M., Mitchell, L. A., Xie, Z. X., Lin, Q. H., Wang, X., Xiao, W. H., Wang, Y., Zhou, X., Liu, H., Li, X., Ding, M. Z., Liu, D., Zhang, L., Liu, B. L., Wu, X. L., Li, F. F., Dong, X. T., ... Yuan, Y. J. (2017). Bug mapping and fitness testing of chemically synthesized chromosome X. Science, 355(6329), Article eaaf4706. https://doi.org/10.1126/science.aaf4706

Wu, Y, Li, BZ, Zhao, M, Mitchell, LA, Xie, ZX, Lin, QH, Wang, X, Xiao, WH, Wang, Y, Zhou, X, Liu, H, Li, X, Ding, MZ, Liu, D, Zhang, L, Liu, BL, Wu, XL, Li, FF, Dong, XT, Jia, B, Zhang, WZ, Jiang, GZ, Liu, Y, Bai, X, Song, TQ, Chen, Y, Zhou, SJ, Zhu, RY, Gao, F, Kuang, Z, Wang, X, Shen, M, Yang, K, Stracquadanio, G, Richardson, SM, Lin, Y, Wang, L, Walker, R, Luo, Y, Ma, PS, Yang, H, Cai, Y, Dai, J, Bader, JS, Boeke, JD & Yuan, YJ 2017, 'Bug mapping and fitness testing of chemically synthesized chromosome X', Science, vol. 355, no. 6329, eaaf4706. https://doi.org/10.1126/science.aaf4706

@article{efd9ea873693406cb897a85cfed14d8f,

title = "Bug mapping and fitness testing of chemically synthesized chromosome X",

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.",

author = "Yi Wu and Li, {Bing Zhi} and Meng Zhao and Mitchell, {Leslie A.} and Xie, {Ze Xiong} and Lin, {Qiu Hui} and Xia Wang and Xiao, {Wen Hai} and Ying Wang and Xiao Zhou and Hong Liu and Xia Li and Ding, {Ming Zhu} and Duo Liu and Lu Zhang and Liu, {Bao Li} and Wu, {Xiao Le} and Li, {Fei Fei} and Dong, {Xiu Tao} and Bin Jia and Zhang, {Wen Zheng} and Jiang, {Guo Zhen} and Yue Liu and Xue Bai and Song, {Tian Qing} and Yan Chen and Zhou, {Si Jie} and Zhu, {Rui Ying} and Feng Gao and Zheng Kuang and Xuya Wang and Michael Shen and Kun Yang and Giovanni Stracquadanio and Richardson, {Sarah M.} and Yicong Lin and Lihui Wang and Roy Walker and Yisha Luo and Ma, {Ping Sheng} and Huanming Yang and Yizhi Cai and Junbiao Dai and Bader, {Joel S.} and Boeke, {Jef D.} and Yuan, {Ying Jin}",

note = "Funding Information: Work in Tianjin University was funded by {"}863{"} program: 2012AA02A708, {"}973{"} program: 2014CB745100, and international cooperative project: 2015DFA00960 from the Ministry of Science and Technology, and China National Natural Science Foundation of China: 21390203 and 21621004. U.S. research was funded by U.S. NSF grants MCB-1026068, MCB-1158201 to J.D.B. and MCB-1445545 to J.S.B., and U.S. Department of Energy DE-FG02097ER25308 to S.M.R.. J.D.B. and J.S.B. are founders and directors of Neochromosome Inc. J.D.B. serves as a scientific advisor to Recombinetics Inc. and Sample6, Inc. These arrangements are reviewed and managed by the committees on Conflict of interest at NYU Langone Medical Center (J.D.B.) and Johns Hopkins University (J.S.B.). U.K. research was funded by a Chancellor's Fellowship from the University of Edinburgh, a start-up fund from Scottish Universities Life Sciences Alliance (SULSA), and Biotechnology and Biological Sciences Research Counci grants BB/M005690/1, BB/M025640/1, and BB/M00029X/1 to Y.C. We thank A. Heguy and the staff at NYU's Genome Technology Center for outstanding deep-sequencing services. Additional information (synX design diagram, PCRTag sequences, Feature summary table (wild-type X, designed synX, physical strain yYW0115; yeast-chr10-9-01), Variants in physical strain (yeast-chr10-9-01), Minichunk plasmids, PCR primers) related to synX can be accessed on the Sc2.0 website http://syntheticyeast.org, under main menu Sc2.0→Data. All genomic data for this paper are available under the Sc2.0 umbrella BioProject accession number PRJNA351844. Requests for materials should be addressed to Y.-J. Yuan ([email protected]). Publisher Copyright: {\textcopyright} 2017, American Association for the Advancement of Science. All rights reserved.",

year = "2017",

month = mar,

day = "10",

doi = "10.1126/science.aaf4706",

language = "English (US)",

volume = "355",

journal = "Science",

issn = "0036-8075",

publisher = "American Association for the Advancement of Science",

number = "6329",

}

TY - JOUR

T1 - Bug mapping and fitness testing of chemically synthesized chromosome X

AU - Wu, Yi

AU - Li, Bing Zhi

AU - Zhao, Meng

AU - Mitchell, Leslie A.

AU - Xie, Ze Xiong

AU - Lin, Qiu Hui

AU - Wang, Xia

AU - Xiao, Wen Hai

AU - Wang, Ying

AU - Zhou, Xiao

AU - Liu, Hong

AU - Li, Xia

AU - Ding, Ming Zhu

AU - Liu, Duo

AU - Zhang, Lu

AU - Liu, Bao Li

AU - Wu, Xiao Le

AU - Li, Fei Fei

AU - Dong, Xiu Tao

AU - Jia, Bin

AU - Zhang, Wen Zheng

AU - Jiang, Guo Zhen

AU - Liu, Yue

AU - Bai, Xue

AU - Song, Tian Qing

AU - Chen, Yan

AU - Zhou, Si Jie

AU - Zhu, Rui Ying

AU - Gao, Feng

AU - Kuang, Zheng

AU - Wang, Xuya

AU - Shen, Michael

AU - Yang, Kun

AU - Stracquadanio, Giovanni

AU - Richardson, Sarah M.

AU - Lin, Yicong

AU - Wang, Lihui

AU - Walker, Roy

AU - Luo, Yisha

AU - Ma, Ping Sheng

AU - Yang, Huanming

AU - Cai, Yizhi

AU - Dai, Junbiao

AU - Bader, Joel S.

AU - Boeke, Jef D.

AU - Yuan, Ying Jin

N1 - Funding Information: Work in Tianjin University was funded by "863" program: 2012AA02A708, "973" program: 2014CB745100, and international cooperative project: 2015DFA00960 from the Ministry of Science and Technology, and China National Natural Science Foundation of China: 21390203 and 21621004. U.S. research was funded by U.S. NSF grants MCB-1026068, MCB-1158201 to J.D.B. and MCB-1445545 to J.S.B., and U.S. Department of Energy DE-FG02097ER25308 to S.M.R.. J.D.B. and J.S.B. are founders and directors of Neochromosome Inc. J.D.B. serves as a scientific advisor to Recombinetics Inc. and Sample6, Inc. These arrangements are reviewed and managed by the committees on Conflict of interest at NYU Langone Medical Center (J.D.B.) and Johns Hopkins University (J.S.B.). U.K. research was funded by a Chancellor's Fellowship from the University of Edinburgh, a start-up fund from Scottish Universities Life Sciences Alliance (SULSA), and Biotechnology and Biological Sciences Research Counci grants BB/M005690/1, BB/M025640/1, and BB/M00029X/1 to Y.C. We thank A. Heguy and the staff at NYU's Genome Technology Center for outstanding deep-sequencing services. Additional information (synX design diagram, PCRTag sequences, Feature summary table (wild-type X, designed synX, physical strain yYW0115; yeast-chr10-9-01), Variants in physical strain (yeast-chr10-9-01), Minichunk plasmids, PCR primers) related to synX can be accessed on the Sc2.0 website http://syntheticyeast.org, under main menu Sc2.0→Data. All genomic data for this paper are available under the Sc2.0 umbrella BioProject accession number PRJNA351844. Requests for materials should be addressed to Y.-J. Yuan ([email protected]). Publisher Copyright: © 2017, American Association for the Advancement of Science. All rights reserved.

PY - 2017/3/10

Y1 - 2017/3/10

N2 - 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.

AB - 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.

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Bug mapping and fitness testing of chemically synthesized chromosome X

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