A single-stage polymerase-based protocol for the introduction of deletions and insertions without subcloning.
A single-stage polymerase-based procedure is described that allows extensive modifications of DNA. The version described here uses the QuikChange Site-Directed Mutagenesis System kit supplied by Stratagene. The original protocol is replaced by a single-stage method in which linear production of complementary strands is accomplished in separate single primer reactions. This has proved effective in introducing insertions and deletions into large gene/vector combinations without subcloning.
Salerno
John C
JC
Biology Department, Rensselaer Polytechnic Institute, Science Center, 1W14, 110 8th St., Troy, NY 12180, USA. salers2@rpi.edu
Jones
Rachel J
RJ
Erdogan
Eda
E
Smith
Susan M E
SM
D017931:DNA Primers / Q000737:chemistry*; D016254:Mutagenesis, Insertional / Q000379:methods; D010957:Plasmids / Q000737:chemistry*; D016133:Polymerase Chain Reaction / Q000379:methods; D017384:Sequence Deletion
D017931:DNA Primers