Figure 2.

Genomic DMS footprinting analysis of the B. japonicum fixRnifA upstream promoter region. The B. japonicum fixRnifA upstream promoter DNA region, integrated on the chromosome as a fixR-lacZ fusion, was analyzed by DMS footprinting in anaerobic cultures of strains A8085 (nifA⁻ background), 8085 (wild-type background), 8091 (mutant A⁻⁶⁸ to C promoter in wild-type background), and N8085 (rpoN1⁻/rpoN2⁻ background). Primer extension products, obtained by linear amplification of the genomic DNA with Taq DNA polymerase, were separated by gel electrophoresis and detected by autoradiography. Guanine residue −121 of the top DNA strand was protected from methylation in all the strains except in the nifA⁻ background, whereas guanine −100 was clearly protected in the 8085 and 8091 strains, and less protected in strain N8085. Guanine residue −75 of the protein binding site (PBS) is also shown (see Fig. 3).