title: Engineering the ribosomal DNA in a megabase synthetic chromosome.
abstract: We designed and synthesized a 976,067-base pair linear chromosome, synXII, based on native chromosome XII inSaccharomyces cerevisiaeSynXII was assembled using a two-step method, specified by successive megachunk integration and meiotic recombination-mediated assembly, producing a functional chromosome inS. cerevisiae.Minor growth defect "bugs" detected in synXII, caused by deletion of tRNA genes, were rescued by introducing an ectopic copy of a single tRNA gene. The ribosomal gene cluster (rDNA) on synXII was left intact during the assembly process and subsequently replaced by a modified rDNA unit used to regenerate rDNA at three distinct chromosomal locations. The signature sequences within rDNA, which can be used to determine species identity, were swapped to generate aSaccharomycessynXII strain that would be identified asSaccharomyces bayanusby standard DNA barcoding procedures.
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.|Tsinghua-Peking Center for Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Weimin
authors.identifier: 0000-0002-8836-1438
authors.initials: W
authors.lastname: Zhang
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.|Peking University-Tsinghua University-National Institute of Biological Sciences Joint Graduate Program, College of Life Science, Peking University, Beijing 100871, China.
authors.forename: Guanghou
authors.identifier: 0000-0002-3192-5423
authors.initials: G
authors.lastname: Zhao
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Zhouqing
authors.identifier: 0000-0001-5426-5176
authors.initials: Z
authors.lastname: Luo
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Yicong
authors.initials: Y
authors.lastname: Lin
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Lihui
authors.identifier: 0000-0002-8479-7859
authors.initials: L
authors.lastname: Wang
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Yakun
authors.identifier: 0000-0002-4847-0648
authors.initials: Y
authors.lastname: Guo
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Ann
authors.initials: A
authors.lastname: Wang
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Shuangying
authors.initials: S
authors.lastname: Jiang
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Qingwen
authors.identifier: 0000-0003-3444-2854
authors.initials: Q
authors.lastname: Jiang
authors.affiliation: BGI-Shenzhen, Shenzhen 518083, China.
authors.forename: Jianhui
authors.identifier: 0000-0002-3910-9260
authors.initials: J
authors.lastname: Gong
authors.affiliation: BGI-Shenzhen, Shenzhen 518083, China.
authors.forename: Yun
authors.identifier: 0000-0001-5872-3269
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Sha
authors.identifier: 0000-0002-4378-0479
authors.initials: S
authors.lastname: Hou
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Jing
authors.initials: J
authors.lastname: Huang
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.|Tsinghua-Peking Center for Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Tianyi
authors.initials: T
authors.lastname: Li
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Yiran
authors.initials: Y
authors.lastname: Qin
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Junkai
authors.initials: J
authors.lastname: Dong
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Qin
authors.initials: Q
authors.lastname: Qin
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.|Tsinghua-Peking Center for Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Jiaying
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Xinzhi
authors.initials: X
authors.lastname: Zou
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Xi
authors.initials: X
authors.lastname: He
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Li
authors.initials: L
authors.lastname: Zhao
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Yibo
authors.initials: Y
authors.lastname: Xiao
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Meng
authors.initials: M
authors.lastname: Xu
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Erchao
authors.identifier: 0000-0001-7829-1870
authors.initials: E
authors.lastname: Cheng
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Ning
authors.identifier: 0000-0001-6857-5386
authors.initials: N
authors.lastname: Huang
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Tong
authors.initials: T
authors.lastname: Zhou
authors.affiliation: BGI-Shenzhen, Shenzhen 518083, China.|School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3BF, UK.|BGI-Qingdao, Qingdao 266555, China.
authors.forename: Yue
authors.initials: Y
authors.lastname: Shen
authors.affiliation: School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3BF, UK.
authors.forename: Roy
authors.identifier: 0000-0003-0960-4591
authors.initials: R
authors.lastname: Walker
authors.affiliation: School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3BF, UK.
authors.forename: Yisha
authors.identifier: 0000-0002-4228-7527
authors.initials: Y
authors.lastname: Luo
authors.affiliation: Institute for Systems Genetics and Department of Biochemistry and Molecular Pharmacology, New York University Langone Medical Center, New York, NY 10011, USA.
authors.forename: Zheng
authors.identifier: 0000-0001-5855-8371
authors.initials: Z
authors.lastname: Kuang
authors.affiliation: Institute for Systems Genetics and Department of Biochemistry and Molecular Pharmacology, New York University Langone Medical Center, New York, NY 10011, USA.
authors.forename: Leslie A
authors.identifier: 0000-0001-7869-9817
authors.initials: LA
authors.lastname: Mitchell
authors.affiliation: High-Throughput Biological Center and Department of Biomedical Engineering, Johns Hopkins University, Baltimore, MD 21205, USA.
authors.forename: Kun
authors.identifier: 0000-0002-1060-8082
authors.initials: K
authors.lastname: Yang
authors.affiliation: High-Throughput Biological Center and Department of Biomedical Engineering, Johns Hopkins University, Baltimore, MD 21205, USA.
authors.forename: Sarah M
authors.identifier: 0000-0002-9820-9421
authors.initials: SM
authors.lastname: Richardson
authors.affiliation: Key Laboratory of Systems Bioengineering (Ministry of Education), SynBio Research Platform, Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), School of Chemical Engineering and Technology, Tianjin University, Tianjin 300072, China.
authors.forename: Yi
authors.identifier: 0000-0002-2539-9228
authors.initials: Y
authors.lastname: Wu
authors.affiliation: Key Laboratory of Systems Bioengineering (Ministry of Education), SynBio Research Platform, Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), School of Chemical Engineering and Technology, Tianjin University, Tianjin 300072, China.
authors.forename: Bing-Zhi
authors.identifier: 0000-0003-4121-3048
authors.initials: BZ
authors.lastname: Li
authors.affiliation: Key Laboratory of Systems Bioengineering (Ministry of Education), SynBio Research Platform, Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), School of Chemical Engineering and Technology, Tianjin University, Tianjin 300072, China.
authors.forename: Ying-Jin
authors.identifier: 0000-0003-0553-0089
authors.initials: YJ
authors.lastname: Yuan
authors.affiliation: BGI-Shenzhen, Shenzhen 518083, China.|James D. Watson Institute of Genome Sciences, Hangzhou 310058, China.
authors.forename: Huanming
authors.identifier: 0000-0003-1703-3012
authors.initials: H
authors.lastname: Yang
authors.affiliation: Wuxi Qinglan Biotechnology Inc., Yixing, Jiangsu 214200, China.
authors.forename: Jiwei
authors.identifier: 0000-0003-3803-6503
authors.initials: J
authors.lastname: Lin
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Guo-Qiang
authors.initials: GQ
authors.lastname: Chen
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Qingyu
authors.initials: Q
authors.lastname: Wu
authors.affiliation: High-Throughput Biological Center and Department of Biomedical Engineering, Johns Hopkins University, Baltimore, MD 21205, USA.
authors.forename: Joel S
authors.identifier: 0000-0002-6020-4625
authors.initials: JS
authors.lastname: Bader
authors.affiliation: School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3BF, UK.
authors.forename: Yizhi
authors.identifier: 0000-0003-1663-2865
authors.initials: Y
authors.lastname: Cai
authors.affiliation: Institute for Systems Genetics and Department of Biochemistry and Molecular Pharmacology, New York University Langone Medical Center, New York, NY 10011, USA.
authors.forename: Jef D
authors.identifier: 0000-0001-5322-4946
authors.initials: JD
authors.lastname: Boeke
authors.affiliation: Key Laboratory for Industrial Biocatalysis (Ministry of Education) and Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China. jbdai@tsinghua.edu.cn.
authors.forename: Junbiao
authors.identifier: 0000-0002-5299-4700
authors.initials: J
authors.lastname: Dai
mesh_terms: D002467:Cell Nucleus / Q000235:genetics / Q000648:ultrastructure; D018244:Chromosomes, Artificial, Yeast / Q000737:chemistry* / Q000235:genetics / Q000648:ultrastructure; D004275:DNA, Ribosomal / Q000235:genetics*; D005818:Genetic Engineering / Q000379:methods*; D016681:Genome, Fungal; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000648:ultrastructure; D058615:Synthetic Biology / Q000379:methods*; D059467:Transcriptome
chemical_list: D004275:DNA, Ribosomal