qss

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P. aeruginosa is an opportunistic human pathogen. It regulates its virulence gene expression implicating QS system. P. aeruginosa contains two LuxI/ LuxR type QS systems, (a) lasI/lasR and, (b) rhII/rhIR. In lasI/lasR system, LasI synthesizes N-3-oxo-dodecanoyl-homoserine lactone (OdDHL), which binds with transcriptional activator LasR, leading to the up-regulation of cognate promoters. In rhII/rhIR system, RhII synthesizes N-butanoyl-homoserine lactone (BHL), which then binds with RhIR, leading to expression of cognate promoters (de Kievit et al., 1999). Pseudomonas based biosensors have been constructed by mutating auto inducer synthase gene (lasI or rhII), and expressing reporter such as lacZ from QS responsive promoter. P. aeruginosa PAO1 M71LZ strain has been used to detect long chain AHL such as C10-oxo- and C12-oxo-AHL. M71LZ is a lasI deletion mutant, and contains chromosomally integrated fusion of rsaL promoter and lacZ (Anbazhagan et al., 2012). Studies have demonstrated that LasI/LasR QS system directly regulates expression of rsaL, making this system a suitable biosensor (de Kievit et al., 1999). Presence of AHL is quantified by measuring the level of lacZ, by performing β-galactosidase assay. Later, a biosensor strain was developed to sense primarily 3-OH-C6-AHL,by engineering PhzI/PhzR QS system of P. fluorescens 2-79 (Khan et al., 2005).PhzI/PhzR QS system regulates the expression of the phzABCDEFG operon. Products of this operon synthesize chorismate and convert it to an antimicrobial compound phenazine-1-carboxylic acid. Autoinducer synthase PhzI synthesizes six different AHLs including, 3-OH-C6-AHL, 3-OH-C8-AHL, 3-OH-C10-AHL. P. fluorescens 2-79 based biosensor carrying engineered PhzI/PhzR QS system is most res... ... middle of paper ... ...nge of AHLs (Zhu et al., 2003). This biosensor was made ultrasensitive by overexpressing traR from T7 promoter. A. tumefaciens strain KYC55, which lacks Ti plasmid and consequently does not produce TraI, was used as a host. Biosensor networks were cloned on 3 plasmids, pJZ372, pJZ384, and pJZ410. Plasmid pJZ384 contains fusion of T7 promoter and traR, plasmid pJZ410 contains T7 RNA polymerase, and plasmid pJZ372 contains PtraI::lacZ fusion, derived from the plasmid pCF372 (Zhu et al., 2003). This strain demonstrated better sensitivity and range compared to a closely related biosensor A. tumefaciens strain WCF47 (pCF218) (pCF372), where traR is expressed from tetracycline inducible promoter (Ptet)(Zhu et al., 1998). Since, this strain uses lacZ reporter gene, can be used with TLC based qualitative, and Miller assay based quantitative assays for the screening of AHLs.

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