A simplified method for generating step-wise deletions using PCR.
A simple and general method is described for the generation of ordered deletions for DNA sequencing. Nicked plasmids, the substrates for step-wise digestion by exonuclease III, are obtained after the self-ligation of PCR products with phosphorylated and non-phosphorylated primers and plasmid DNAs as template. The method is suitable for use with any plasmid vector and for generation of deletion clones with deletions in both possible directions.
Kitabatake
M
M
Department of Biophysics, Faculty of Science, Kyoto University, Japan.
Inokuchi
H
H
D001483:Base Sequence; D003001:Cloning, Molecular; D004277:DNA, Single-Stranded; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D010766:Phosphorylation; D010957:Plasmids / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods*; D017422:Sequence Analysis, DNA / Q000379:methods*; D017384:Sequence Deletion
D004277:DNA, Single-Stranded; D009838:Oligodeoxyribonucleotides