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. 2002 Dec;184(24):6929–6941. doi: 10.1128/JB.184.24.6929-6941.2002

FIG. 1.

FIG. 1.

Identification of a repetitive sequence in the genome of M. fermentans PG18. (A) Genomic DNA from M. fermentans PG18 (clone 39) was digested with BamHI, and the resulting fragments were separated by standard agarose electrophoresis. Five BamHI fragments, corresponding to visible ethidium bromide-stained bands in the 3- to 5-kb size range, were cloned, and the nucleotide sequences were determined. The size of each fragment (calculated from the nucleotide sequence) is indicated in kilobases. The locations and directions of ORFs within each fragment are shown by open arrows, and the ORFs are labeled with the standard gene names for ORFs that are housekeeping genes with known functions. ORF1 to ORF12 correspond to ORF1 to ORF12 in Fig. 2. ORF488 and ORF299 are designated by length (in amino acid residues) and have significant homology (BLAST P < 1E-16) to conserved hypothetical ORFs MG443 (from M. genitalium) and Mypu4350 (from M. pulmonis), respectively. The asterisk represents the location of the hybridization probe (primer 2; see Materials and Methods) used in the Southern analysis in panel B. Some terminal ORFs indicated by arrows (for example, orf5) are incomplete by virtue of the BamHI restriction site by which their fragments were cloned. (B) Southern hybridization analysis of multiple copies of ICEF in M. fermentans PG18. Genomic DNA was digested with BssHII (lane 1), BglII (lane 2), BglII and BssHII (lane 3), BamHI (lane 4), or XbaI (lane 5) and probed with DIG-labeled oligonucleotide primer 2 (see Materials and Methods). Lane 6 contained DIG-labeled λ HindIII markers (Roche), the sizes of which are indicated in kilobase pairs at the right of panel C. The open triangle highlights a reproducible but weakly hybridizing BssHII restriction fragment of approximately 5.7 kb that was present in lane 1. (C) Southern analysis of multiple copies of the right terminal portion of ICEF. Genomic DNA from M. fermentans PG18 was digested with MluI (lane 1), BssHII (lane 2), MluI-BssHII (lane 3), or XbaI (lane 4) and probed with DIG-labeled primer 7 (see Materials and Methods and Fig. 2). Lane 5 contained DIG-labeled markers as in panel B.