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Megaprimer-Based PCR to Synthesize Fusion Genes for Cloning

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 2967))

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Abstract

Megaprimer-based polymerase chain reaction (PCR) strategies allow the versatile and fast assembly and amplification of a myriad of tailor-made or random DNA sequences readily available for conventional or restriction-free (RF) cloning.

In this chapter, we present a megaprimer-based PCR protocol that enables the expeditious construction of customized fusion genes ready for cloning into commercial expression plasmids. With the expanding use of protein tag technology in the most diverse application fields, this protocol remains a versatile and affordable solution for the synthesis and fusion of peptide tags/domains of interest.

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References

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Acknowledgments

This study was supported by the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UIDB/04469/2020 unit and project ESSEntial (PTDC/BII-BTI/1858/2021).

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Author notes

  1. Tatiana Q. Aguiar and Carla Oliveira contributed equally to this work.

Authors and Affiliations

  1. CEB - Centre of Biological Engineering, University of Minho, Braga, Portugal

    Tatiana Q. Aguiar & Lucília Domingues

  2. LABBELS - Associate Laboratory, Braga/Guimarães, Portugal

    Tatiana Q. Aguiar & Lucília Domingues

  3. Universidade Católica Portuguesa, CBQF - Centro de Biotecnologia e Química Fina – Laboratório Associado, Escola Superior de Biotecnologia, Rua Diogo Botelho, Porto, Portugal

    Carla Oliveira

Authors
  1. Tatiana Q. Aguiar
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  2. Carla Oliveira
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  3. Lucília Domingues
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Corresponding author

Correspondence to Tatiana Q. Aguiar .

Editor information

Editors and Affiliations

  1. CEB - Centre of Biological Engineering, University of Minho, Braga, Portugal

    Lucília Domingues

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© 2023 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature

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Aguiar, T.Q., Oliveira, C., Domingues, L. (2023). Megaprimer-Based PCR to Synthesize Fusion Genes for Cloning. In: Domingues, L. (eds) PCR. Methods in Molecular Biology, vol 2967. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3358-8_16

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  • DOI: https://doi.org/10.1007/978-1-0716-3358-8_16

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-3357-1

  • Online ISBN: 978-1-0716-3358-8

  • eBook Packages: Springer Protocols

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