The rpoN gene cluster and the mutation schemes. (a) Genes surrounding rpoN are shown as open arrows. HP: conserved hypothetical protein with unknown function; YhbG: ABC transporter, ATP binding protein; YfiA: ribosomal subunit interface-associated sigma-54 modulation protein; HprK: Hpr(Ser) kinase/phosphorylase. Insertion of a kanamycin resistance cassette upstream or downstream of the intergenic region of the rpoN gene resulted in viable mutants (a). (b) Scheme showing attempts of construction of deletion of (i) the 5'-end, (ii) the whole, or (iii) the 3'-end of the rpoN coding region. (c) An extra copy of the rpoN gene was inserted on the chromosome and was under the control of the chloramphenicol resistance cassette promoter. (d) An extra copy of the rpoN gene was introduced in trans under the control of a lac promoter (constitutively expressed) or a taclac promoter (IPTG-inducible). The position of insertion of the antibiotic resistance cassette (kanamycin, Kan or gentamycin, Gm) is indicated with an inverted triangle and a vertical bar. The regions which were attempted to replace with the antibiotic resistance cassette insertion are indicated by dashed line.