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j5 DNA Assembly Design Automation

Part of the Methods in Molecular Biology book series (MIMB,volume 1116)

Abstract

Modern standardized methodologies, described in detail in the previous chapters of this book, have enabled the software-automated design of optimized DNA construction protocols. This chapter describes how to design (combinatorial) scar-less DNA assembly protocols using the web-based software j5. j5 assists biomedical and biotechnological researchers construct DNA by automating the design of optimized protocols for flanking homology sequence as well as type IIS endonuclease-mediated DNA assembly methodologies. Unlike any other software tool available today, j5 designs scar-less combinatorial DNA assembly protocols, performs a cost–benefit analysis to identify which portions of an assembly process would be less expensive to outsource to a DNA synthesis service provider, and designs hierarchical DNA assembly strategies to mitigate anticipated poor assembly junction sequence performance. Software integrated with j5 add significant value to the j5 design process through graphical user-interface enhancement and downstream liquid-handling robotic laboratory automation.

Key words

  • DNA assembly
  • Design automation
  • BioCAD
  • Combinatorial library
  • Synthetic biology

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References

  1. Ellis T, Adie T, Baldwin GS (2011) DNA assembly for synthetic biology: from parts to pathways and beyond. Integr Biol (Camb) 3:109–118. doi:10.1039/c0ib00070a

    CAS  CrossRef  Google Scholar 

  2. Hillson NJ (2011) DNA assembly method standardization for synthetic biomolecular circuits and systems. In: Koeppl H, Densmore D, di Bernardo M, Setti G (eds) Design and analysis of bio-molecular circuits, 1st edn. Springer, New York, pp 295–314

    CrossRef  Google Scholar 

  3. Densmore D, Hsiau TH, Kittleson JT et al (2010) Algorithms for automated DNA assembly. Nucleic Acids Res 38:2607–2616. doi:10.1093/nar/gkq165

    CAS  PubMed Central  PubMed  CrossRef  Google Scholar 

  4. Shetty RP, Endy D, Knight TF Jr (2008) Engineering BioBrick vectors from BioBrick parts. J Biol Eng 2:5. doi:10.1186/1754-1611-2-5

    PubMed Central  PubMed  CrossRef  Google Scholar 

  5. Anderson JC, Dueber JE, Leguia M et al (2010) BglBricks: a flexible standard for biological part assembly. J Biol Eng 4:1. doi:10.1186/1754-1611-4-1

    PubMed Central  PubMed  CrossRef  Google Scholar 

  6. Leguia M, Brophy J, Densmore D et al (2011) Automated assembly of standard biological parts. Methods Enzymol 498:363–397. doi:10.1016/B978-0-12-385120-8.00016-4

    CAS  PubMed  Google Scholar 

  7. Beal J, Weiss R, Densmore D et al (2012) An end-to-end workflow for engineering of biological networks from high-level specifications. ACS Synth Biol 1:317. doi:10.1021/sb300030d

    CAS  PubMed  CrossRef  Google Scholar 

  8. Weber E, Engler C, Gruetzner R et al (2011) A modular cloning system for standardized assembly of multigene constructs. PLoS One 6:e16765. doi:10.1371/journal.pone.0016765

    CAS  PubMed Central  PubMed  CrossRef  Google Scholar 

  9. Sarrion-Perdigones A, Falconi EE, Zandalinas SI et al (2011) GoldenBraid: an iterative cloning system for standardized assembly of reusable genetic modules. PLoS One 6:e21622. doi:10.1371/journal.pone.0021622

    CAS  PubMed Central  PubMed  CrossRef  Google Scholar 

  10. Quan J, Tian J (2009) Circular polymerase extension cloning of complex gene libraries and pathways. PLoS One 4:e6441. doi:10.1371/journal.pone.0006441

    PubMed Central  PubMed  CrossRef  Google Scholar 

  11. Shao Z, Luo Y, Zhao H (2011) Rapid characterization and engineering of natural product biosynthetic pathways via DNA assembler. Mol Biosyst 7:1056–1059. doi:10.1039/c0mb00338g

    CAS  PubMed  CrossRef  Google Scholar 

  12. Carothers JM, Goler JA, Juminaga D et al (2011) Model-driven engineering of RNA devices to quantitatively program gene expression. Science 334:1716–1719. doi:10.1126/science.1212209

    CAS  PubMed  CrossRef  Google Scholar 

  13. Salis HM, Mirsky EA, Voigt CA (2009) Automated design of synthetic ribosome binding sites to control protein expression. Nat Biotechnol 27:946–950. doi:10.1038/nbt.1568

    CAS  PubMed Central  PubMed  CrossRef  Google Scholar 

  14. Egbert RG, Klavins E (2012) Fine-tuning gene networks using simple sequence repeats. Proc Natl Acad Sci U S A 109:16817–16822. doi:10.1073/pnas.1205693109

    CAS  PubMed Central  PubMed  CrossRef  Google Scholar 

  15. Mutalik VK, Guimaraes JC, Cambray G et al (2013) Quantitative estimation of activity and quality for collections of functional genetic elements. Nat Methods 10(4):347–353. doi:10.1038/nmeth.2403

  16. Mutalik VK, Guimaraes JC, Cambray G et al (2013) Precise and reliable gene expression via standard transcription and translation initiation elements. Nat Methods 10(4):354 –360

    Google Scholar 

  17. Hillson NJ, Rosengarten RD, Keasling JD (2012) j5 DNA assembly design automation software. ACS Synth Biol 1:14–21. doi:10.1021/Sb2000116

    CAS  PubMed  CrossRef  Google Scholar 

  18. Chen J, Densmore D, Ham TS et al (2012) DeviceEditor visual biological CAD canvas. J Biol Eng 6:1. doi:10.1186/1754-1611-6-1

    PubMed Central  PubMed  Google Scholar 

  19. Ham TS, Dmytriv Z, Plahar H et al (2012) Design, implementation and practice of JBEI-ICE: an open source biological part registry platform and tools. Nucleic Acids Res 40:e141. doi:10.1093/nar/gks531

    PubMed Central  PubMed  CrossRef  Google Scholar 

  20. Linshiz G, Stawski N, Poust S et al (2012) PaR-PaR laboratory automation platform. ACS Synth Biol 2:216–222. doi:10.1021/sb300075t

    PubMed  CrossRef  Google Scholar 

  21. Thieme F, Engler C, Kandzia R et al (2011) Quick and clean cloning: a ligation-independent cloning strategy for selective cloning of specific PCR products from non-specific mixes. PLoS One 6:e20556. doi:10.1371/journal.pone.0020556

    CAS  PubMed Central  PubMed  CrossRef  Google Scholar 

  22. Li MZ, Elledge SJ (2007) Harnessing homologous recombination in vitro to generate recombinant DNA via SLIC. Nat Methods 4:251–256. doi:10.1038/nmeth1010

    CAS  PubMed  CrossRef  Google Scholar 

  23. You C, Zhang XZ, Zhang YH (2012) Simple cloning via direct transformation of PCR product (DNA Multimer) to Escherichia coli and Bacillus subtilis. Appl Environ Microbiol 78:1593–1595. doi:10.1128/AEM.07105-11

    CAS  PubMed Central  PubMed  CrossRef  Google Scholar 

  24. Quan J, Tian J (2011) Circular polymerase extension cloning for high-throughput cloning of complex and combinatorial DNA libraries. Nat Protoc 6:242–251. doi:10.1038/nprot.2010.181

    CAS  PubMed  CrossRef  Google Scholar 

  25. Erijman A, Dantes A, Bernheim R et al (2011) Transfer-PCR (TPCR): a highway for DNA cloning and protein engineering. J Struct Biol 175:171–177. doi:10.1016/j.jsb.2011.04.005

    CAS  PubMed  CrossRef  Google Scholar 

  26. Zhang Y, Werling U, Edelmann W (2012) SLiCE: a novel bacterial cell extract-based DNA cloning method. Nucleic Acids Res 40:e55. doi:10.1093/nar/gkr1288

    CAS  PubMed Central  PubMed  CrossRef  Google Scholar 

  27. Gibson DG, Young L, Chuang RY et al (2009) Enzymatic assembly of DNA molecules up to several hundred kilobases. Nat Methods 6:343–345. doi:10.1038/nmeth.1318

    CAS  PubMed  CrossRef  Google Scholar 

  28. Ramon A, Smith HO (2011) Single-step linker-based combinatorial assembly of promoter and gene cassettes for pathway engineering. Biotechnol Lett 33:549–555. doi:10.1007/s10529-010-0455-x

    CAS  PubMed  CrossRef  Google Scholar 

  29. Shao Z, Zhao H, Zhao H (2009) DNA assembler, an in vivo genetic method for rapid construction of biochemical pathways. Nucleic Acids Res 37:e16. doi:10.1093/nar/gkn991

    PubMed Central  PubMed  CrossRef  Google Scholar 

  30. Wingler LM, Cornish VW (2011) Reiterative recombination for the in vivo assembly of libraries of multigene pathways. Proc Natl Acad Sci U S A 108:15135–15140. doi:10.1073/pnas.1100507108

    PubMed Central  PubMed  CrossRef  Google Scholar 

  31. j5 website. http://j5.jbei.org

  32. Gibthon website. http://gibthon.org

  33. Richardson SM, Liu S, Boeke JD et al (2012) Design-A-Gene with GeneDesign. Methods Mol Biol 852:235–247. doi:10.1007/978-1-61779-564-0_18

    CAS  PubMed  CrossRef  Google Scholar 

  34. Richardson SM, Nunley PW, Yarrington RM et al (2010) GeneDesign 3.0 is an updated synthetic biology toolkit. Nucleic Acids Res 38:2603–2606. doi:10.1093/nar/gkq143

    CAS  PubMed Central  PubMed  CrossRef  Google Scholar 

  35. GeneDesign website. http://54.235.254.95/gd/

  36. Convert PCR Primers Into In-Fusion® Primers website. http://www.clontech.com/US/Products/Cloning_and_Competent_Cells/Cloning_Kits/xxclt_onlineToolsLoad.jsp?citemId=http://bioinfo.clontech.com/infusion/convertPcrPrimersInit.do%26xxheight=750

  37. GeneArt® Primer and Construct Design Tool website. http://bioinfo.invitrogen.com/oligoDesigner

  38. Engler C, Gruetzner R, Kandzia R et al (2009) Golden gate shuffling: a one-pot DNA shuffling method based on type IIS restriction enzymes. PLoS One 4:e5553. doi:10.1371/journal.pone.0005553

    PubMed Central  PubMed  CrossRef  Google Scholar 

  39. Engler C, Kandzia R, Marillonnet S (2008) A one pot, one step, precision cloning method with high throughput capability. PLoS One 3:e3647. doi:10.1371/journal.pone.0003647

    PubMed Central  PubMed  CrossRef  Google Scholar 

  40. Engler C, Marillonnet S (2011) Generation of families of construct variants using golden gate shuffling. Methods Mol Biol 729:167–181. doi:10.1007/978-1-61779-065-2_11

    PubMed  CrossRef  Google Scholar 

  41. Geertsma ER, Dutzler R (2011) A versatile and efficient high-throughput cloning tool for structural biology. Biochemistry 50:3272–3278. doi:10.1021/bi200178z

    CAS  PubMed  CrossRef  Google Scholar 

  42. Bitinaite J, Rubino M, Varma KH et al (2007) USER friendly DNA engineering and cloning method by uracil excision. Nucleic Acids Res 35:1992. doi:10.1093/nar/gkm041

    CAS  PubMed Central  PubMed  CrossRef  Google Scholar 

  43. Annaluru N, Muller H, Ramalingam S (2012) Assembling DNA fragments by USER fusion. Methods Mol Biol 852:77–95. doi:10.1007/978-1-61779-564-0_7

    CAS  PubMed  CrossRef  Google Scholar 

  44. Olsen LR, Hansen NB, Bonde MT et al (2011) PHUSER (Primer Help for USER): a novel tool for USER fusion primer design. Nucleic Acids Res 39:W61–W67. doi:10.1093/nar/gkr394

    CAS  PubMed Central  PubMed  CrossRef  Google Scholar 

  45. PHUSER website. http://www.cbs.dtu.dk/services/phuser/

  46. TeselaGen website. http://teselagen.com

  47. VectorEditor stand-alone software. https://public-registry.jbei.org/static/vesa/VectorEditor.html

  48. Public instance of the JBEI Parts Registry. http://public-registry.jbei.org

  49. VectorEditor Project. http://code.google.com/p/vectoreditor/

  50. PaR-PaR website. http://prpr.jbei.org

  51. PaR-PaR source code github repository. https://github.com/jbei/prpr

  52. Synthetic Biology Open Language visual standard. http://www.sbolstandard.org/visual

  53. j5 user’s manual. http://j5.jbei.org/j5manual/index.html

  54. Eugene website. http://eugenecad.org/

  55. Bilitchenko L, Liu A, Cheung S et al (2011) Eugene—a domain specific language for specifying and constraining synthetic biological parts, devices, and systems. PLoS One 6:e18882. doi:10.1371/journal.pone.0018882

    CAS  PubMed Central  PubMed  CrossRef  Google Scholar 

  56. Rozen S, Skaletsky H (2000) Primer3 on the WWW for general users and for biologist programmers. Methods Mol Biol 132:365–386

    CAS  PubMed  Google Scholar 

  57. DINAMelt web server. http://mfold.rna.albany.edu/?q=DINAMelt/Quickfold

  58. j5, DeviceEditor, and VectorEditor demonstration video. http://j5.jbei.org/j5_and_DeviceEditor_Demo_Movie.mov

  59. DeviceEditor user’s manual. http://j5.jbei.org/DeviceEditor_manual/index.html

  60. A plasmid Editor (ApE) software. http://biologylabs.utah.edu/jorgensen/wayned/ape/

  61. j5 web-form interface. http://j5.jbei.org/bin/j5_entry_form.pl

  62. Clotho website. http://clothocad.org

  63. Xia B, Bhatia S, Bubenheim B (2011) Developer’s and user’s guide to Clotho v2.0 A software platform for the creation of synthetic biological systems. Methods Enzymol 498:97–135. doi:10.1016/B978-0-12-385120-8.00005-X

    CAS  PubMed  Google Scholar 

  64. DeviceEditor software. http://j5.jbei.org/bin/deviceeditor.pl

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Acknowledgments

Conflict of Interest Statement: The author declares competing financial interests in the form of pending patent applications related to the j5 software, and equity in TeselaGen Biotechnology, Inc. This work conducted by the Joint BioEnergy Institute and the U.S. Department of Energy Joint Genome Institute was supported by the Office of Science, Office of Biological and Environmental Research, of the U.S. Department of Energy under Contract No. DE-AC02-05CH11231. The author thanks Joanna Chen for constructive comments on the manuscript.

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Hillson, N.J. (2014). j5 DNA Assembly Design Automation. In: Valla, S., Lale, R. (eds) DNA Cloning and Assembly Methods. Methods in Molecular Biology, vol 1116. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-764-8_17

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  • DOI: https://doi.org/10.1007/978-1-62703-764-8_17

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  • Publisher Name: Humana Press, Totowa, NJ

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