Biology Reference
In-Depth Information
volume of Tris-HCl (pH 8.0) saturated phenol was added and
proteins were precipitated by mixing with cold methanol contain-
ing 0.1 M ammonium acetate. Using this optimized protocol,
authors successfully constructed soybean leaf proteome map of
high quality with more than 500 reproducibly detected protein
spots in CBB-stained gel. All these observations indicate that both
phenol and TCA/acetone based extraction buffers are suitable for
protein extraction from different tissues of soybean. However, as
compared to TCA/acetone method, the protein yield is compara-
tively higher in phenol-based method in particular for soybean
seed proteins.
3
Comprehensive Analysis of Soybean Proteins
In spite of the diffi culties in extracting proteins from soybean, sub-
stantial soybean proteomic research has been carried out at differ-
ent developmental stages in particular seed maturation and young
seedling stages.
3.1 Seed Maturation
Stage
Seed fi lling is the developmental period when rapid metabolic and
morphological (seed size, weight and color) changes take place
[
25
]. To better understand the metabolic processes associated with
seed fi lling in soybean, Agrawal et al. [
26
] investigated the seed
proteome at fi ve developmental stages by two complementary pro-
teomic approaches—2-DE and semicontinuous multidimensional
protein identifi cation technology (Sec-MudPIT) coupled with liq-
uid chromatography-MS. In total, 478 nonredundant proteins
were collectively identifi ed through 2-DE and Sec-MudPIT analy-
ses, of which only 70 proteins were common to both datasets.
Major functional classes of proteins revealed from both 2-DE and
Sec-MudPIT analyses were involved in primary metabolism, pro-
tein destination and storage, and energy. Sec-MudPIT approach
identifi ed threefold higher number of membrane proteins than
2-DE. Comparisons of quantitative seed-fi lling proteome of soy-
bean and rapeseed were done to further understand the regulation
of intermediary metabolism in protein-rich versus oil-rich seeds.
Authors conclude that 2-DE and Sec-MudPIT could be used as
complementary proteomic approaches for characterizing the soy-
bean seed-fi lling proteome.
Similar proteomic study was previously performed by Hajduch
et al. [
23
] to determine the expression profi le of soybean seed pro-
teins at 2, 3, 4, 5, and 6 weeks after fl owering. Using 2-DE and
matrix-assisted laser desorption ionization time-of-fl ight (MALDI)
MS, high-resolution proteome reference maps, expression profi les
of 679 spots, and corresponding MALDI MS spectra for each spot
were successfully established. Searching NCBI soybean UniGene
database using the MS-Fit program of Protein Prospector, led to
