A single-stage polymerase-based protocol for the introduction of deletions and insertions without subcloning

Molecular Biotechnology

Volumn 29, Issue 3, 2005, Pages 225-232

  Salerno, John C ^a   Jones, Rachel J ^a   Erdogan, Eda ^a   Smith, Susan M E ^a  

^a Research Rensselaer Polytechnic Institute   (United States)

Author keywords

Linear amplification; Site directed mutagenesis

Indexed keywords

CHEMICAL REACTIONS; CLONING; DNA; GENES; MUTAGENESIS;

LINEAR AMPLIFICATION; SITE-DIRECTED MUTAGENESIS; SUBCLONING; VECTOR COMBINATIONS;

GENETIC ENGINEERING;

ARTICLE; DNA MODIFICATION; GENE DELETION; GENE INSERTION; MOLECULAR CLONING; MUTAGENESIS; POLYMERASE CHAIN REACTION; SITE DIRECTED MUTAGENESIS;

DNA PRIMERS; MUTAGENESIS, INSERTIONAL; PLASMIDS; POLYMERASE CHAIN REACTION; SEQUENCE DELETION;

EID: 26244438851     PISSN: 10736085     EISSN: None     Source Type: Journal    
DOI: 10.1385/MB:29:3:225     Document Type: Article

References (9)

1. Mullis, K., Faloona, F., Scharf, S., Saiki, R., Horn, G., and Ehrlich, H. (1986) Specific enzymatic amplification of DNA in vitro: the polymerase chain reaction. Cold Spring Harbor Symp. Quant. Biol. 51, 262-273.
2. Cormack, B. (1994) Mutagenesis of cloned DNA. In: Current Protocols in Molecular Biology (Ausubel, F. M., Brent, R. E., Kingston, D. D., et al., eds.), John Wiley and Sons, New York, pp. 8.5.1-8.5.12.
3. Sarkar, G. and Sommer, S. S. (1990) The "megaprimer" method of site-directed mutagenesis. Biotechniques 8, 404-407.
4. Aiyar, A. and Leis, J. (1993) Modification of the megaprimer method of PCR mutagenesis: improved amplificaiton of the final product. Biotechniques 15, 366-369.
5. Barik, S. (1996) Site-directed mutagenesis in virto by megaprimer PCR. Methods Mol. Biol. 57, 203-215.
6. Wang, W. Y. and Malcolm, B. A. (1999) Two-stage PCR protocol allowing introduction of multiple mutations, deletions and insertions using QuikChange Site-Directed Mutagenesis. Biotechniques 26, 680-682.
7. Wang, W. Y. and Malcolm, B. A. (2002)Two-stage polymerase chain reaction protocol allowing introduction of multiple mutations, deletions, and insertions, using QuikChange site-directed mutagenesis. Methods Mol. Biol. 182, 37-43.
8. Makarova, O., Kamberov, E., and Margolis, B. (2000) Generation of deletion and point mutations with one primer in a single cloning. Biotechniques 29, 970-972.
9. Geiser, M., Cèbe, R., Drewello, D., and Schmitz, R. (2001) Integration of PCR fragments at any specific site within cloning vectors without the use of restriction enzymes and DNA ligase Biotechniques 31, 88-92.