A proteomic study of pseudomonas putida by two-dimensional gel electrophoresis: Establishing quantitative standards for intra-laboratory results

Two-dimensional gel electrophoresis (2DE) is the most commonly employed technique in the field of proteomics. The technique is designed to separate thousands proteins in highly complex biological samples. It is largely because of the diversity of the sample that 2DE has encountered problems with reproducibility. This study was designed to address and alleviate many of these problems and also to establish quantitative standards for intra-laboratory results. A universal standard gel of Pseudomonas putida KT2440 was created and tested. The universal standard gel will serve as an intra-laboratory reference to which future experiments will be compared. An acceptable coefficient of variation between future control gels and the universal standard gel was determined to be less than 35%. Another aim of this study is to explore the effects of growth time on protein expression and determine a window of time in which growth can be stopped and protein expression is preserved. Currently there have been no studies relating to this topic. Protein samples were harvested every 30 minutes throughout the log-phase of growth and their protein expression was determined. It was found that within a 30 minute window of time surrounding mid-log phase the protein expression does not change, within statistical limits. This study will serve as the quantitative cornerstone for all future intra-laboratory experiments