[The application of two-dimensional liquid phase chromatography to separation of total protein of Leymus chinensis shoot].

Abstract

The total proteins of Leymus chinensis were extracted from eight-week-old seedlings grown in the greenhouse by TCA-acetone precipitation method. After the lysis buffer was replaced by the start buffer, proteins were fractionated along the first dimension using chromatofocusing (CF). Subsequently the fractions with pI values between 8.5 and 4.0 collected after the first dimension separation were further fractionated by nonporous silica reverse-phase high-performance liquid chromatography (NPS-RP-HPLC, HPRP). With ProteoVue software the pI/UV map was generated to show the protein expression profile of Leymus chinensis. Some experiments were tested to optimize the fraction procedure. Three different elution gradients were employed to get the optimal chromatogram. Comparison of the protein expression pattern between 2D-PAGE and 2D-LC indicated that 2D-LC was a powerful tool in protein fraction. Reproducibility and veracity of the protein patterns were confirmed in different injections of the same sample. A method to fractionate the total protein of Leymus chinensis shoot with two-dimensional liquid chromatography (2D-LC) was founded.