Characterization of plant growth-promoting rhizobacteria using capillary isoelectric focusing with whole column imaging detection.
Capillary isoelectric focusing (cIEF) can be a useful tool for the characterization and identification of microbes. Based on the whole column imaging detection (WCID) technique and using plant growth-promoting rhizobacteria (PGPR) as test microbes, we present a two-level cIEF characterization method for the characterization and identification of bacteria. Intact bacteria were first characterized according to their apparent isoelectric points measured by cIEF-WCID and then lysed bacteria were further characterized by cIEF profiling of the intracellular proteins. Cellular clustering was found to be the main experimental barrier for the characterization of intact bacteria. The addition of sodium chloride (100mM) to the sample mixture was found to be an effective way to reduce clustering. Due to the high efficiency and high resolution of cIEF-WCID, characterization of bacteria according to their intracellular proteins can be implemented simply and quickly without optimization of the experimental conditions. To improve the detection sensitivity with laser induced fluorescence (LIF)-WCID, the possibility to label bacteria with a non-covalent fluorescent dye, NanoOrange, was explored.
Liu
Zhen
Z
Department of Chemistry, Nanjing University, Nanjing 210093, China. zhenliu@nju.edu.cn
Wu
Shan Shan
SS
Pawliszyn
Janusz
J
D001426:Bacterial Proteins / Q000302:isolation & purification; D019075:Electrophoresis, Capillary / Q000379:methods*; D016972:Enterobacter cloacae / Q000302:isolation & purification; D020563:Gammaproteobacteria / Q000302:isolation & purification*; D007525:Isoelectric Focusing / Q000379:methods*; D009930:Organic Chemicals; D011549:Pseudomonas / Q000302:isolation & purification; D013010:Sonication
D001426:Bacterial Proteins; C433144:NanoOrange; D009930:Organic Chemicals