Extraction of Total Protein from Axillary Buds of Sugarcane (Saccharum spp.) for Proteomic Analysis

Abstract

Since sugar cane production depends on the germination of axillary buds, the analysis of the products of gene expression in axillary buds from different sugarcane varieties (Saccharum spp.) in each growth cycle is important to reveal the proteins related to sprouting potential for crop renovation and consequent increase in sugarcane production. Current study has employed a simple method for protein extraction from axillary buds of RB867515, RB866928 and CTC04 sugarcane varieties to obtain protein fractions for two-dimensional electrophoresis and proteomic analysis. The TCA/acetone method for protein extraction with modifications in the centrifugation processes and in the incubation time of samples revealed a large number of well-defined bands in SDS-PAGE system with molecular weights ranging between 120 and 10 kDa. The evidence that the method TCA/acetone modified (1) by using 200 mg of tissue axillary buds; (2) by inclusion of an incubation step at room temperature for 20 min under stirring; (3) by aliquots division for a better recovery from the supernatant; (4) by the absence of phenol in one extraction phase; and (5) by the increase in centrifugal rotation from 12,000×g to 13,860×g and an increase from 15 to 30 min centrifugation time (in the precipitation and wash steps) was more efficient than the other methods used for sugarcane, is one more example showing that methods for extracting proteins in plants are specific to different tissues.