Expression of quorum sensing genes
We performed gene assays to evaluate the impact of chitosan on the
expression of QS-regulated lasR and rhlR genes. The P. aeruginosaisolate was cultivated in LB medium with or without chitosan at a
concentration of 100 μg /ml. RNA was isolated using TRIzol reagent
(Sigma-Aldrich, UK) and dissolved in 20 μl of 0.1%
diethylpyrocarbonate
(DEPC)-treated water. On the same day, cDNA was amplified using cDNA
Synthesis kit (Yekta Tajhiz Azma Co, Iran) according to the
manufacturer’s instructions. The
quantitative polymerase chain
reaction (qPCR) reactions were conducted using the 7500 Sequence
Detection System (Applied Biosystems Inc., Foster, CA, USA) and Power
SYBR Green PCR Master Mix (Applied Biosystems). Gene-specific primers
were adopted from (Muslim et al., 2018a) listed in Table 3 were employed
to determine the expression level of QS-regulated genes. The rpsL gene
was utilized as a control to normalize the expression of lasR and rhlR
genes, and the experiments were carried out in triplicate for real-time
analysis.