Abstract
Restriction digest analysis and Sanger sequencing are among the most commonly used techniques to check the sequence of synthetic DNA constructs. However, both require careful preparation to select restriction enzymes or DNA primers adapted to the expected constructs sequences. In projects involving manufacturing of large batches of synthetic constructs, the task can be tedious and error-prone. This chapter demonstrates the use of two free and open-source web applications providing fast and automated selection of enzymes and sequencing primers for DNA construct verification.
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References
Casini A, Storch M, Baldwin GS, Ellis T (2015) Bricks and blueprints: methods and standards for DNA assembly. Nat Rev Mol Cell Biol 16(9):568–576. https://doi.org/10.1038/nrm4014
Potapov V, Ong JL, Kucera RB, Langhorst BW, Bilotti K, Pryor JM et al (2018) Comprehensive profiling of four base overhang ligation fidelity by T4 DNA ligase and application to DNA assembly. ACS Synth Biol 7(11):2665–2674. https://doi.org/10.1021/acssynbio.8b00333
Shapland EB, Holmes V, Reeves CD, Sorokin E, Durot M, Platt D et al (2015) Low-cost, high-throughput sequencing of DNA assemblies using a highly multiplexed Nextera process. ACS Synth Biol 4(7):860–866. https://doi.org/10.1021/sb500362n
Sanger F, Coulson AR (1975) A rapid method for determining sequences in DNA by primed synthesis with DNA polymerase. J Mol Biol 94(3):441–448. https://doi.org/10.1016/0022-2836(75)90213-2
Dharmadi Y, Patel K, Shapland E, Hollis D, Slaby T, Klinkner N et al (2014) High-throughput, cost-effective verification of structural DNA assembly. Nucleic Acids Res 42(4):e22. https://doi.org/10.1093/nar/gkt1088
Hancock JM, Zvelebil MJ, Hancock JM (2004). PRIMER3. In: Dictionary of bioinformatics and computational biology. https://doi.org/10.1002/9780471650126.dob0560.pub2
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Zulkower, V. (2021). Computer-Aided Planning for the Verification of Large Batches of DNA Constructs. In: Menolascina, F. (eds) Synthetic Gene Circuits . Methods in Molecular Biology, vol 2229. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1032-9_7
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DOI: https://doi.org/10.1007/978-1-0716-1032-9_7
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