A simple and efficient method for the oligonucleotide-directed mutagenesis using plasmid DNA template and phosphorothioate-modified nucleotide

Manabu Sugimoto, Nobuyoshi Esaki, Hidehiko Tanaka, Kenji Soda

Research output: Contribution to journalArticle

31 Citations (Scopus)

Abstract

We have developed a simple and efficient method for oligonucleotide-directed mutagenesis with double-stranded (plasmid) DNA as a template. The template was simply and rapidly prepared by cell lysis and the following DNA denaturation with alkali. The chain elongation was performed with phosphorothioate-modified nucleotide at 37°C. After the selective digestion of original DNA with NciI and exonuclease III, the desired mutated gene was obtained at a high frequency (about 70%).

Original languageEnglish
Pages (from-to)309-311
Number of pages3
JournalAnalytical Biochemistry
Volume179
Issue number2
DOIs
Publication statusPublished - 1989
Externally publishedYes

Fingerprint

Mutagenesis
Site-Directed Mutagenesis
Oligonucleotides
Plasmids
Nucleotides
Nucleic Acid Denaturation
DNA
Alkalies
Digestion
Denaturation
Elongation
Genes
exodeoxyribonuclease III

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology

Cite this

A simple and efficient method for the oligonucleotide-directed mutagenesis using plasmid DNA template and phosphorothioate-modified nucleotide. / Sugimoto, Manabu; Esaki, Nobuyoshi; Tanaka, Hidehiko; Soda, Kenji.

In: Analytical Biochemistry, Vol. 179, No. 2, 1989, p. 309-311.

Research output: Contribution to journalArticle

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