## 2 Results
### Construction of a Strain with a Modified lapA Transcription and a LapA Strain without a vWFa Domain  
Although we knew that Fis increases _lapA_ transcription and LapA is required for biofilm formation (Figures 1 and 3), we had to split the Fis-effect from the LapA-effect on the biofilm. There are two reasons: (i) Fis can enhance biofilm through another factor than LapA, and (ii) _P. putida_ forms the biofilm even in the absence of LapA in the cell [1; 20]. Therefore, _P. putida_ strains were constructed in which native _lapA_ is under the P\({}_{\mathit{tuc}}\) promoter, and IPTG controls its transcription. _P. putida_ strain PaW85 (isogenic for KT2440), one of the standard strains of _P. putida_, was used for this purpose (Table 1). Additionally, the PSm derivative of the PaW85 was used to evaluate the biofilm in the context of PSm and F15 (Table 1). The construction of the _lapA_ expression system was problematic for several reasons. Firstly, _lapA_ is a very long gene (26 kbp) [12]; therefore, we could not use vector systems. Secondly, chromosomal _lapA_ shares a promoter region with its transporter genes _lapBC_ encoded opposite to _lapA_[12; 20]. The disruption of the joint promoter region can affect the transcription of both genes. We used several expression systems to control the transcription of both _lapA_ and _lapBC_ genes. For example, an expression system was constructed that ensured the expression of _lapA_ from the P\({}_{\mathit{tuc}}\) promoter by IPTG and the transcription of _lapBC_ from the P\({}_{\mathit{m}}\) promoter by benzoate (data not shown). In this case, transcription in both directions was regulated by extracellularly added effectors. Unfortunately, the addition of phenolic compounds to the medium affected the biofilm formation per se; these strains were excluded from the study (data not shown). We had to use a construct that had the P\({}_{\mathit{tuc}}\) promoter immediately upstream of the lapA gene. The reversely expressed _lapBC_ retained the entire native promoter area without the possibility of external regulation by effectors. With this system, we constructed a P-PANB strain from PaW85 (Table 1). This variant is not ideal because, despite induced _lapA_ transcription, the effect on the biofilm may remain modest, as the natively expressed transporter may begin to limit LapA exposure on the outer membrane. The third part of the LapA transporter, LapE, is described in _P. fluorescens_ as an outer membrane subunit where LapA N-terminus will trap. In this way, the extracellular LapA is anchored in the outer membrane [32]. The _lapE_ deletion prevents the LapA transport to the outside of the cell, and the biofilm should weaken. Thus, this deletant strain was used as a negative control for P-PANB strains. The _lapG_ deletion, on the other hand, was used to construct a positive control. LapG is a periplasmic protease that, when activated, cleaves the N-terminus of LapA and releases LapA into the environment [32], resulting in a reduction of biofilm. Thus, the deletion of _lapG_ should produce a robust _P. putida_ biofilm, as LapA is not removed from the outer membrane. Before this research, we had constructed _lapE_ and _lapG_ deletion variants from _P. putida_ strain PSm, PSm-E, and PSm-G, respectively. Therefore, we created the IPTG-inducible _lapA_ transcription strains PSm-E-PANB and PSm-G-PANB, based on PSm-E and PSm-G, respectively (Table 1). The sequence coding a von Willebrand factor type A (vWFa) domain in _P. putida lapA_ was deleted (from amino acid S8167 to 8281P, designated Avwf\({}^{-}\)). Recent studies have shown that the deletion of the vWFa domain in _P. fluorescens_ LapA reduces biofilm formation to glass and hydrophilic plastic [10; 32]. The von Willebrand factor itself, after which the bacterial adhesin domains were named, is a multimeric glycoprotein important for the protein-protein interaction that causes thrombus formation and bleeding arrest after injury of eukaryotic tissue [15]. Inspired by these works, we asked whether the vWFa domain of _P.