Abstract
Mutagenesis by the overlap extension PCR has become a standard method of creating mutations including substitutions, insertions, and deletions. Nonetheless, the established overlap PCR mutagenesis is limited in many respects. In particular, it has been difficult to make an insertion larger than 30 nt, since all sequence alterations must be embedded within the primer. Here, we describe a rapid and efficient method for creating insertions or deletions of any length at any position in a DNA molecule. This method is generally applicable, and therefore represents a significant improvement to the now widely used overlap extension PCR method.
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Acknowledgments
We thank Dr. Byung-Yoon Ahn (Korea University) for critical reading of the manuscript. This work was supported by a grant R01-2006-000-10620-0 from the Basic Research Program of the Korea Science & Engineering Foundation, Republic of Korea.
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Lee, J., Shin, MK., Ryu, DK., Kim, S., Ryu, WS. (2010). Insertion and Deletion Mutagenesis by Overlap Extension PCR. In: Braman, J. (eds) In Vitro Mutagenesis Protocols. Methods in Molecular Biology, vol 634. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-60761-652-8_10
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DOI: https://doi.org/10.1007/978-1-60761-652-8_10
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