Abstract
Recombinant PCR (1) is the method of choice if one wants to modify a cloned DNA. It is a versatile technique that allows operations as different as creation of deletions, addition of small insertions, site directed mutagenesis, and construction of chimeric molecules at any chosen location in the molecule of interest (see Note 1). This chapter describes in detail a simplification of the original recombinant PCR method. This fast and efficient method has been successful in fusing two different sequences with precision (2–4). It can also be used to create deletions or insert small fragments of DNA.
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References
Higuchi, R. (1989) Using PCR to engineer DNA, in PCR Technology, Principles and Applications for DNA Amplification. Stockton, New York, pp. 61–70.
Pont-Kingdon, G. (1994) Construction of chimeric molecules by a two-step recombinant PCR method. Biotechniques 16, 1010,1011.
Cao, Y. (1990) Direct cloning of a chimeric gene fused by the polymerase chain reaction. Technique 2, 109–111.
Yon, J. and Fried, M. (1989) Precise gene fusion by PCR. Nucleic Acids Res. 17, 4895.
Sharrocks, A. D. (1994) The design of primers for PCR, in PCR Technology Current Innovations (Griffin, G. G. and Griffin A. M., eds.), CRC, Boca Raton, FL, pp. 5–11.
Klug, J., Wolf, M., and Beato, M. (1991) Creating chimeric molecules by PCR directed homologous DNA recombination. Nucleic Acids Res. 19, 2793.
Sarkar, G. and Sommer, S. S. (1990) The “megaprimer” method of site-directed mutagenesis. BioTechniques 8, 404–407.
Barik, S. (1993) Site-directed mutagenesis by double polymerase chain reaction, in Methods in Molecular Biology, vol. 15: PCR Protocols: Current Methods and Applications (White, B. A., ed.), Humana, Totowa, NJ, pp. 277–286.
Saiki, R. K. (1989) The design and optimization of the PCR, in PCR Technology. Principles and Applications for DNA Amplification (Erlich, H. A., ed.), Stockton, New York, pp. 7–16.
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© 1997 Humana Press Inc.
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Pont-Kingdon, G. (1997). Creation of Chimeric Junctions, Deletions, and Insertions by PCR. In: White, B.A. (eds) PCR Cloning Protocols. Methods in Molecular Biology™, vol 67. Humana Press, Totowa, NJ. https://doi.org/10.1385/0-89603-483-6:167
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DOI: https://doi.org/10.1385/0-89603-483-6:167
Publisher Name: Humana Press, Totowa, NJ
Print ISBN: 978-0-89603-483-9
Online ISBN: 978-1-59259-553-2
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