Abstract:

Objective　To set up a rapid, efficient, reliable and accurate method for separation ofBacteriodesforsythuspro- teins.Methods　BacteroidesforsythusATCC43037 cells were harvested by centrifugation, washed inTris buffer to remove excess medium, and lysed by sonication. The sonicated lysis proteins were extracted step by step with ReadyPrepTMSequential Extraction Kit (Bio-Rad). The supernatant ofB.forsythusproteins were used for two-dimensional gel electrophoresis. The first dimension IEF(isoelectric focusing) was run with Immobiline DryStrip (pH 3~10) and the second dimension SDS-PAGE was run with Ex- celgel SDS,gradient 8-18 precast gel and buffer strips. The separated proteinswere stainedwith Coomassie Brilliant Blue and silver staining kit (Plusone Silver Staining Kit, Protein, Pharmacia Biotech).Results　1. The protein spots were clear when sample cups were used in the middle of IPG strip during IEF. 2.B.forsythusproteinswere separated clearly by horizontal two-dimension- al electrophoresis and most ofB.forsythuswhole-cell proteins were acidic proteins (PI3-7).Conclusion　Horizontal two-dimen- sion electrophoresis is a useful method for separatingB.forsythusproteins.

Key words: horizontal two-dimensional electrophoresis, Bacteriodesforsythus, protein separation