A GENETIC SYSTEM FOR ARCHAEA OF THE GENUS METHANOSARCINA - LIPOSOME-MEDIATED TRANSFORMATION AND CONSTRUCTION OF SHUTTLE VECTORS

New methods that allow, for the first time, genetic analysis in Archae a of the genus Methanosarcina are presented, First, several autonomous ly replicating plasmid shuttle vectors have been constructed based ore the naturally occurring plasmid pC2A from Methanosarcina acetivorans. These vectors replicate in 9 of 11 Methanosarcina strains tested and in Escherichia coli. Second, a highly efficient transformation system based upon introduction of DNA by liposomes has been developed, This m ethod allows transformation frequencies of as high as 2 x 10(8) transf ormants per microgram of DNA per 10(9) cells or approximate to 20% of the recipient population. During the course of this work, the complete 5467-bp DNA sequence of pC2A was determined. The implications of thes e findings for the future of methanoarchaeal research are also discuss ed.