Electrotransformation of Streptococcus pneumoniae: evidence for restriction of DNA on entry.
Electrotransformation is a method generally used in biotechnology to introduce recombinant DNA into a wide range of bacteria. However, the mechanism of DNA entry is poorly understood. We report that in Streptococcus pneumoniae, a naturally transformable species, electrotransformation efficiently introduces a plasmid replicon. DNA is strongly restricted by the restriction-modification systems DpnI and DpnII which degrade methylated and non-methylated DNA, respectively, at GATC sequences. This suggests that in electrotransformation double-stranded DNA penetrates into these bacteria without a single-stranded DNA step in contrast to natural transformation. Single-stranded DNA by itself is able to electrotransform very weakly and linearized double-stranded plasmid DNA yields barely detectable levels of transformants.
Lefrançois
Jacques
J
Laboratoire de Microbiologie et Génétique Moléculaire du CNRS and UniversitéPaul Sabatier, 118 route de Narbonne, 31062 Toulouse Cedex, France.
Sicard
A Michel
AM
Laboratoire de Microbiologie et Génétique Moléculaire du CNRS and UniversitéPaul Sabatier, 118 route de Narbonne, 31062 Toulouse Cedex, France.
D019175:DNA Methylation; D015280:DNA Restriction-Modification Enzymes; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D018274:Electroporation; D010957:Plasmids / Q000235:genetics; D013296:Streptococcus pneumoniae / Q000235:genetics*; D014170:Transformation, Genetic
D015280:DNA Restriction-Modification Enzymes; C020102:endodeoxyribonuclease DpnI; D015252:Deoxyribonucleases, Type II Site-Specific; C024405:GATC-specific type II deoxyribonucleases