Skip to main content
SpringerLink
Log in

Construction of Minimal Genomes and Synthetic Cells

  • Chapter
  • First Online:
Minimal Cells: Design, Construction, Biotechnological Applications

  • 1006 Accesses

Abstract

A minimal genome strain containing only genes necessary for maintaining self-replicable life was proposed as a potential platform having various advantages in chemical and pharmaceutical industries. With recent advances in high-throughput DNA sequencing and synthesis technology, many reduced genomes have now been constructed. In this chapter, we will review previously constructed artificially reduced genomes to confirm the potential of their industrial utility. Some of them exhibit growth rates similar to those of their parental wild-type strains while offering higher genetic stability and productivity. Furthermore, we will discuss some technological hurdles and limitations encountered during the design and construction of reduced genomes.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Chapter
USD 29.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 149.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 199.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 199.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

Abbreviations

ALE:

Adaptive laboratory evolution

asRNA:

Antisense RNA

CDS:

Coding sequence

COG:

Clusters of orthologous genes

CRISPR:

Clustered regularly interspaced short palindromic repeats

IS element:

Insertion sequence element

kbp:

Kilo base pair

LUCA:

Last universal common ancestor

Mbp:

Mega base pair

ORF:

Open reading frame

RNAi:

RNA interference

sgRNA:

Single guide RNA

References

  • Ara K, Ozaki K, Nakamura K et al (2007) Bacillus minimum genome factory: effective utilization of microbial genome information. Biotechnol Appl Biochem 46:169–178

    Article  CAS  PubMed  Google Scholar 

  • Baba T, Ara T, Hasegawa M et al (2006) Construction of Escherichia coli K-12 in-frame, single-gene knockout mutants: the Keio collection. Mol Syst Biol 2(2006):0008

    PubMed  Google Scholar 

  • Bennett GM, Moran NA (2013) Small, smaller, smallest: the origins and evolution of ancient dual symbioses in a phloem-feeding insect. Genome Biol Evol 5:1675–1688

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Biswas I, Gruss A, Ehrlich SD, Maguin E (1993) High-efficiency gene inactivation and replacement system for gram-positive bacteria. J Bacteriol 175:3628–3635

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Brown JR, Douady CJ, Italia MJ, Marshall WE, Stanhope MJ (2001) Universal trees based on large combined protein sequence data sets. Nat Genet 28:281–285

    Article  CAS  PubMed  Google Scholar 

  • Charlebois RL, Doolittle WF (2004) Computing prokaryotic gene ubiquity: rescuing the core from extinction. Genome Res 14:2469–2477

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Choe D, Lee JH, Yoo M et al (2019) Adaptive laboratory evolution of a genome-reduced Escherichia coli. Nat Commun 10:935

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Christen B, Abeliuk E, Collier JM et al (2011) The essential genome of a bacterium. Mol Syst Biol 7:528

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Gerdes SY, Scholle MD, Campbell JW et al (2003) Experimental determination and system level analysis of essential genes in Escherichia coli MG1655. J Bacteriol 185:5673–5684

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Ghatak S, King ZA, Sastry A, Palsson BO (2019) The y-ome defines the 35% of Escherichia coli genes that lack experimental evidence of function. Nucleic Acids Res 47:2446–2454

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Gibson DG, Glass JI, Lartigue C et al (2010) Creation of a bacterial cell controlled by a chemically synthesized genome. Science 329:52–56

    Article  CAS  PubMed  Google Scholar 

  • Giga-Hama Y, Tohda H, Takegawa K, Kumagai H (2007) Schizosaccharomyces pombe minimum genome factory. Biotechnol Appl Biochem 46:147–155

    Article  CAS  PubMed  Google Scholar 

  • Glass JI, Assad-Garcia N, Alperovich N et al (2006) Essential genes of a minimal bacterium. Proc Natl Acad Sci U S A 103:425–430

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Harris JK, Kelley ST, Spiegelman GB, Pace NR (2003) The genetic core of the universal ancestor. Genome Res 13:407–412

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Hartleb D, Jarre F, Lercher MJ (2016) Improved metabolic models for E. coli and Mycoplasma genitalium from GlobalFit, an algorithm yhat simultaneously matches growth and non-growth data sets. PLoS Comput Biol 12:e1005036

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Hashimoto M, Ichimura T, Mizoguchi H et al (2005) Cell size and nucleoid organization of engineered Escherichia coli cells with a reduced genome. Mol Microbiol 55:137–149

    Article  CAS  PubMed  Google Scholar 

  • Hefzi H, Ang KS, Hanscho M et al (2016) A consensus genome-scale reconstruction of chinese hamster ovary cell metabolism. Cell Syst 3:434–443 e438

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Hirashima K, Iwaki T, Takegawa K, Giga-Hama Y, Tohda H (2006) A simple and effective chromosome modification method for large-scale deletion of genome sequences and identification of essential genes in fission yeast. Nucleic Acids Res 34:e11

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Hutchison CA 3rd, Chuang RY, Noskov VN et al (2016) Design and synthesis of a minimal bacterial genome. Science 351:aad6253

    Article  PubMed  CAS  Google Scholar 

  • Hutchison CA, Peterson SN, Gill SR et al (1999) Global transposon mutagenesis and a minimal Mycoplasma genome. Science 286:2165–2169

    Article  CAS  PubMed  Google Scholar 

  • Itaya M (1995) An estimation of minimal genome size required for life. FEBS Lett 362:257–260

    Article  CAS  PubMed  Google Scholar 

  • Ji Y, Zhang B, Van SF et al (2001) Identification of critical staphylococcal genes using conditional phenotypes generated by antisense RNA. Science 293:2266–2269

    Article  CAS  PubMed  Google Scholar 

  • Kobayashi K, Ehrlich SD, Albertini A et al (2003) Essential Bacillus subtilis genes. Proc Natl Acad Sci U S A 100:4678–4683

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Kolisnychenko V, Plunkett G 3rd, Herring CD et al (2002) Engineering a reduced Escherichia coli genome. Genome Res 12:640–647

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Komatsu M, Uchiyama T, Omura S, Cane DE, Ikeda H (2010) Genome-minimized Streptomyces host for the heterologous expression of secondary metabolism. Proc Natl Acad Sci U S A 107:2646–2651

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Koonin EV (2003) Comparative genomics, minimal gene-sets and the last universal common ancestor. Nat Rev Microbiol 1:127–136

    Article  CAS  PubMed  Google Scholar 

  • Lajoie MJ, Rovner AJ, Goodman DB et al (2013) Genomically recoded organisms expand biological functions. Science 342:357–360

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Lee JH, Sung BH, Kim MS et al (2009) Metabolic engineering of a reduced-genome strain of Escherichia coli for L-threonine production. Microb Cell Factories 8:2

    Article  CAS  Google Scholar 

  • Lin DC, Grossman AD (1998) Identification and characterization of a bacterial chromosome partitioning site. Cell 92:675–685

    Article  CAS  PubMed  Google Scholar 

  • Lluch-Senar M, Delgado J, Chen WH et al (2015) Defining a minimal cell: essentiality of small ORFs and ncRNAs in a genome-reduced bacterium. Mol Syst Biol 11:780

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • McCutcheon JP, Moran NA (2011) Extreme genome reduction in symbiotic bacteria. Nat Rev Microbiol 10:13–26

    Article  PubMed  CAS  Google Scholar 

  • Mizoguchi H, Sawano Y, Kato J, Mori H (2008) Superpositioning of deletions promotes growth of Escherichia coli with a reduced genome. DNA Res 15:277–284

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Moran NA, Mira A (2001) The process of genome shrinkage in the obligate symbiont Buchnera aphidicola. Genome Biol 2:RESEARCH0054

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Morimoto T, Kadoya R, Endo K et al (2008) Enhanced recombinant protein productivity by genome reduction in Bacillus subtilis. DNA Res 15:73–81

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Moya A, Gil R, Latorre A et al (2009) Toward minimal bacterial cells: evolution vs. design. FEMS Microbiol Rev 33:225–235

    Article  CAS  PubMed  Google Scholar 

  • Mushegian AR, Koonin EV (1996) A minimal gene set for cellular life derived by comparison of complete bacterial genomes. Proc Natl Acad Sci U S A 93:10268–10273

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Nijman SM (2011) Synthetic lethality: general principles, utility and detection using genetic screens in human cells. FEBS Lett 585:1–6

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Nolan RP, Lee K (2011) Dynamic model of CHO cell metabolism. Metab Eng 13:108–124

    Article  CAS  PubMed  Google Scholar 

  • Orth JD, Conrad TM, Na J et al (2011) A comprehensive genome-scale reconstruction of Escherichia coli metabolism--2011. Mol Syst Biol 7:535

    Article  PubMed  PubMed Central  Google Scholar 

  • Park MK, Lee SH, Yang KS et al (2014) Enhancing recombinant protein production with an Escherichia coli host strain lacking insertion sequences. Appl Microbiol Biotechnol 98:6701–6713

    Article  CAS  PubMed  Google Scholar 

  • Plesa C, Sidore AM, Lubock NB, Zhang D, Kosuri S (2018) Multiplexed gene synthesis in emulsions for exploring protein functional landscapes. Science 359:343–347

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Pósfai G, Plunkett G 3rd, Feher T et al (2006) Emergent properties of reduced-genome Escherichia coli. Science 312:1044–1046

    Article  CAS  PubMed  Google Scholar 

  • Qi LS, Larson MH, Gilbert LA et al (2013) Repurposing CRISPR as an RNA-guided platform for sequence-specific control of gene expression. Cell 152:1173–1183

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Quan J, Saaem I, Tang N et al (2011) Parallel on-chip gene synthesis and application to optimization of protein expression. Nat Biotechnol 29:449–452

    Article  CAS  PubMed  Google Scholar 

  • Reuß DR, Altenbuchner J, Mader U et al (2017) Large-scale reduction of the Bacillus subtilis genome: consequences for the transcriptional network, resource allocation, and metabolism. Genome Res 27:289–299

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Richardson SM, Mitchell LA, Stracquadanio G et al (2017) Design of a synthetic yeast genome. Science 355:1040–1044

    Article  CAS  PubMed  Google Scholar 

  • Riley M, Abe T, Arnaud MB et al (2006) Escherichia coli K-12: a cooperatively developed annotation snapshot--2005. Nucleic Acids Res 34:1–9

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Rousset F, Cui L, Siouve E et al (2018) Genome-wide CRISPR-dCas9 screens in E. coli identify essential genes and phage host factors. PLoS Genet 14:e1007749

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Shalem O, Sanjana NE, Hartenian E et al (2014) Genome-scale CRISPR-Cas9 knockout screening in human cells. Science 343:84–87

    Article  CAS  PubMed  Google Scholar 

  • Sleator RD (2010) The story of Mycoplasma mycoides JCVI-syn1.0: the forty million dollar microbe. Bioeng Bugs 1:229–230

    PubMed  PubMed Central  Google Scholar 

  • Srinivasan R, Scolari VF, Lagomarsino MC, Seshasayee AS (2015) The genome-scale interplay amongst xenogene silencing, stress response and chromosome architecture in Escherichia coli. Nucleic Acids Res 43:295–308

    Article  CAS  PubMed  Google Scholar 

  • Wang T, Wei JJ, Sabatini DM, Lander ES (2014) Genetic screens in human cells using the CRISPR-Cas9 system. Science 343:80–84

    Article  CAS  PubMed  Google Scholar 

  • Weaver DS, Keseler IM, Mackie A, Paulsen IT, Karp PD (2014) A genome-scale metabolic flux model of Escherichia coli K-12 derived from the EcoCyc database. BMC Syst Biol 8:79

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Westers H, Dorenbos R, van Dijl JM et al (2003) Genome engineering reveals large dispensable regions in Bacillus subtilis. Mol Biol Evol 20:2076–2090

    Article  CAS  PubMed  Google Scholar 

  • Yu BJ, Sung BH, Koob MD et al (2002) Minimization of the Escherichia coli genome using a Tn5-targeted Cre/loxP excision system. Nat Biotechnol 20:1018–1023

    Article  CAS  PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Biological Science, Korea Advanced Institute of Science and Technology, Daejeon, South Korea

    Donghui Choe, Sun Chang Kim & Byung-Kwan Cho

  2. KI for the BioCentury, Korea Advanced Institute of Science and Technology, Daejeon, South Korea

    Donghui Choe, Sun Chang Kim & Byung-Kwan Cho

  3. Intelligent Synthetic Biology Center, Daejeon, South Korea

    Sun Chang Kim & Byung-Kwan Cho

  4. Department of Bioengineering, University of California, San Diego, CA, USA

    Bernhard O. Palsson

  5. Bioinformatics and Systems Biology Program, University of California, San Diego, CA, USA

    Bernhard O. Palsson

  6. Department of Pediatrics, University of California, San Diego, CA, USA

    Bernhard O. Palsson

  7. Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Lyngby, Denmark

    Bernhard O. Palsson

Authors
  1. Donghui Choe
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Sun Chang Kim
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Bernhard O. Palsson
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Byung-Kwan Cho
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Byung-Kwan Cho .

Editor information

Editors and Affiliations

  1. Departamento de Procesos y Tecnología, Universidad Autónoma Metropolitana-Cuajimalpa, Ciudad de Mexico, Mexico

    Alvaro R. Lara

  2. Departamento de Ingeniería Celular y Biocatálisis, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Cuernavaca, Morelos, Mexico

    Guillermo Gosset

Rights and permissions

Reprints and permissions

Copyright information

© 2020 Springer Nature Switzerland AG

About this chapter

Check for updates. Verify currency and authenticity via CrossMark

Cite this chapter

Choe, D., Kim, S.C., Palsson, B.O., Cho, BK. (2020). Construction of Minimal Genomes and Synthetic Cells. In: Lara, A., Gosset, G. (eds) Minimal Cells: Design, Construction, Biotechnological Applications. Springer, Cham. https://doi.org/10.1007/978-3-030-31897-0_2

Download citation

Publish with us

Policies and ethics

Search

Navigation