Two-codon insertion mutagenesis of plasmid genes by using single-stranded hexameric oligonucleotides Academic Article uri icon

Overview

MeSH Major

  • Genetic Engineering
  • Mutation
  • Oligodeoxyribonucleotides
  • Oligonucleotides

abstract

  • An efficient method for introducing two codons into a cloned gene has been applied to studying functional regions of the pBR322-encoded tetracycline-resistance gene and beta-lactamase (ampicillin-resistance) gene. Single-stranded hexameric linkers are inserted into a preexisting cohesive end restriction site to create a new (six-base recognition) restriction site. Insertion mutations are enriched by using biochemical selection or are selected by using a kanamycin-resistance cassette (biological selection). Phenotypes of insertion mutations isolated in the tetracycline-resistance gene support the hypothesis that it is comprised of two domains connected by a central hinge. Mutations in the beta-lactamase gene are temperature sensitive and demonstrate altered sensitivity to various beta-lactams and inhibitors.

publication date

  • January 1985

Research

keywords

  • Academic Article

Identity

Language

  • eng

PubMed Central ID

  • PMC397964

PubMed ID

  • 3889924

Additional Document Info

start page

  • 4202

end page

  • 6

volume

  • 82

number

  • 12