## 2 Results
### The Transcription of lapA Is Growth Medium Independent, the Amount of LapA Is Not  
Because the absence of LapA abolished the peptide-dependence effect on the _P. putida_ biofilm (Figure 1), the question arose whether the peptide dependency on the _P. putida_ biofilm could be caused via _lapA_ expression. Accordingly, the _lapA_ expression in _P. putida_ was assessed at transcriptional and protein levels. The _lapA_ transcription was evaluated by \(\beta\)-galactosidase assay, and the abundance of LapA protein was estimated in crude cell lysates (Figure 3). LacZ activity was measured using plasmid p9_PlapA1-8, which carries the _lapA_ promoter region in front of the reporter gene _lacZ_ (Table 1). Figure 3: The expression of adhesins LapA and LapF in _P. putida_ strains PSm and F15. (**A**) \(\beta\)-galactosidase activity in cells carrying pB_PlapA1-8 where _lapA_ promoter region cloned in front of _lacZ_ gene. Arithmetical means of at least three independent sets of measurements and 95% confidence intervals of the arithmetic mean. The homogenous groups are shown above columns by lower-case letters. Identical letters denote nonsignificant differences (\(p\geq 0.05\)) among averages of means. (**B**) SDS-PAA gel electrophoresis from cell lysates prepared from cells grown in M9-0.2 CAA, M9-0.2 CAA+tryptone or LB medium with or without IPTG. PSm\(\Delta\)lapA and PSm\(\Delta\)lapF cell lysates were loaded onto the gel as negative controls of LapA and LapF. The arrows indicate the location of LapA and LapF. 40 mg of total protein was applied to the line. (**C**) Normalised LapA and (**D**) LapF amount. The amount of adhesine determined from the crude lysate of cells grown with IPTG was divided by the amount of LapA determined from the crude lysate of cells grown without IPTG. The averages of at least 6 biological parallels are given, with 95% confidence intervals. The homogenous groups are shown above columns by lower-case letters. Identical letters denote nonsignificant differences (\(p\geq 0.05\)) between the averages of the ratios. Our results revealed that transcription of _lapA_ was dependent on the medium but did not correlate with the F15 biofilm (Figures 1 and 3A) [19]. F15 revealed the positive effect of _fs_-overexpression only in the presence of peptides (Figure 1). In all media, LacZ activity in F15 in the presence of 1 mM IPTG was 1.26 (M9-0.2 CAA), 1.3 (M9-0.2 CAA+tryptone), and 1.48 (LB) times higher than in the non-IPTG variant (_p_ < 0.0001), indicating that regardless of the growth medium, the _lapA_ transcription was always increased in the presence of the higher amount of Fis (Figure 3A). LacZ activity in PSm or F15 containing the empty vector p9 TTBlacZ was less than 0.5 Miller units. \begin{table}
\begin{tabular}{c c c} \hline \hline
**Strain or Plasmid** & **Genotype or Description** & **Reference** \\ \hline _E. coli_ & & \\ CC118 \(\lambda\)pir & \(\Delta\)(_ara-leu_) _araD \(\Delta\)lacX74 gLE galle  
The crude cell lysates were prepared from the bacteria grown for 18 h in the appropriate medium to quantify the adhesins LapA and LapF. Because cell lysates prepared from different media were electrophoresed on PAA gels separately, the ratio of adhesins with and without IPTG was compared separately in every medium. In contrast to the level of _lapA_ transcription, the amount of LapA protein in the crude cell lysate was affected by the content of the growth medium of bacteria (Figure 3B-D). Since the effect of Fis on the F15 biofilm was evident (Figures 1 and 2), we focused on comparing the amount of LapA in F15. Overexpression of _fis_ in the tryptone-containing medium increased the amount of LapA 1.50 times (M9-0.2 CAA) and 1.46 times (LB) in the F15 strain, which correlated with the previously observed increase in F15 biofilm formation in the same growth conditions (Figures 1 and 3C) [19]. However, the _fis_-overexpression did not elevate the amount of LapA in F15 grown in the M9-0.2 CAA medium (Figure 3C).