Bacterial artificial chromosome (BAC) library as a tool for physical mapping of the archaeon Methanosarcina...

We have used a variety of methods to characterize the genome of the archaeon Methanosarcina thermophila TM-1. Pulsed-field gel analysis indicates a genome size of 2.8 Mb. We have constructed a bacterial artificial chromosome (BAC) library of M. thermophila and have used it to generate physical maps for this organism. The library is made up of 384 clones with an average insert size of 58 kb representing 8.0 genome equivalents. The utility of the library for low-resolution physical mapping was shown by identifying NotI linking clones and using these to order the NotI macrorestriction fragments of M. thermophila into a 2.8 Mb map. Hybridization of nine single copy genes and a 16S rRNA sequence to these macrorestriction fragments forms the basis for the first genetic map in this organism. High-resolution physical maps, consisting of overlapping clones, have been created using HindIII fingerprints of BAC clones. In this way, we identified a minimal path of five clones that span a 270 kb NotI fragment. The ease of manipulating BAC clones makes the BAC system an excellent choice for the construction of low-resolution and high-resolution physical and genetic maps of archaeal genomes. It also provides a substrate for future genome-sequencing efforts.