Transformation ofMethanococcus maripaludisand identification of aPstI-like restriction system

An optimized polyethylene glycol (PEG) method of transformation was developed for Methanococcus maripaludis using the pKAS102 integration vector. The frequency of transformation with 0.8 μg of plasmid and 3×109 cells was 4.8×10−5 transformants cfu−1, or 1.8×105 transformants μg−1, which was four orders of magnitude greater than with the natural transformation method. A PstI restriction activity in M. maripaludis was also identified. Methylation of the plasmid with PstI methylase increased the methanococcal transformation frequency at least four-fold. Also, chromosomal DNA from M. maripaludis was resistant to digestion by the PstI endonuclease.