Figure 4. The Sinorhizobium fredii greA and lpsB genes are not cotranscribed but greA affects lpsB expression.

A, genetic organization of the greA-lpsB-lpsE region. Solid lines show the position of the greA (1, 284-bp) and lpsB (3, 156-bp) internal fragments that were successfully amplified (positive controls) using cDNA or gDNA as template. Dotted lines show the positions of the PCR fragments covering the intergenic greA-lpsB (2, 156-bp) and lpsB-lpsE (4, 282-bp) regions that were amplified from gDNA but not from cDNA. B, agarose gel electrophoresis of samples resulting from PCR and RT-PCR experiments. lanes 1–4, controls without DNA; lane 5, 100bp-ladder DNA marker; lanes 6–9, HH103 cDNA; lanes 10–13, HH103 gDNA. Primers used: greAintF and greAintR (lanes 1, 6, and 10), greAlpsB-F and greAlpsB-R (lanes 2, 7, and 11), rtlpsB-F and rtlpsB-R (lanes 3, 8, and 12), lpsBE-F and lpsBE-R (lanes 4, 9, and 13). DNA marker 100, 200, and 300-bp bands are indicated on the left of the figure. C, relative lpsB expression of S. fredii HH103 strains SVQ655 (greA::lacZΔp-Gm^R, →→) and SVQ656 (greA::lacZΔp-Gm^R, →←) in comparison to that of wild type HH103 Rif^R. RNA was extracted from early exponential phase cultures grown in YMB. Each strain was pairwise compared with HH103 Rif^R by using the Mann-Whitney non-parametrical test. In both cases, the relative expression of lpsB was significantly different from that of HH103 Rif^R at the levels α = 5% (indicated with a black asterisk),