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Submitted to: BMC Genomics
Publication Type: Peer Reviewed Journal Publication Acceptance Date: 12/8/2016 Publication Date: 1/6/2017 Citation: Mahalingam, R. 2017. Shotgun proteomics of the barley seed proteome. Biomed Central Genomics. 18:44. Interpretive Summary: Barely seeds are the primary raw materials for the brewing industry. The amount and composition of the barely seed proteins are key factors that determine the suitability and quality of the grain for its end use. In this research a state-of-the art technique for in-depth analysis of the seed proteome was used to identify more than 1110 proteins. Comparative analysis of a two-rowed and six-rowed barley cultivar indicated significant differences in 20 seed proteins that included the three hordoindoline proteins. Phenotypic differences in the seed hardness trait may be associated with the differences in the hordoindolines. The top-down proteomics strategy used in this research can be used for the analysis of more complex traits such as malting quality to identify novel protein markers useful for the brewing industry. Technical Abstract: Barley seed proteins are of prime importance to the brewing industry, human and animal nutrition and in plant breeding for cultivar identification. To obtain comprehensive proteomic data from barley seeds, acetone precipitated proteins were in-solution digested and the resulting peptides were analyzed by nano-liquid chromatography coupled with tandem mass spectrometry. Protein isolations and mass spectrometry were conducted on three biological replicate samples from a two-rowed (Conrad) and a six-rowed (Lacey) barley cultivar. The raw mass spectra data searched against Uniprot’s Barley database using in-house Mascot search engine identified 1168 unique proteins. Gene Ontology (GO) analysis indicated that the majority of the seed proteins were cytosolic, with catalytic activity and associated with carbohydrate metabolism. GO enrichment analysis using the orthologous rice proteins were undertaken using AgriGO database. Spectral counting analysis showed that there are 20 differentially expressed seed proteins between the two-rowed Conrad and six-rowed Lacey cultivars. Phenotypic differences in the seed hardness trait in these two cultivars may be associated with the differential expression of hordoindoline proteins. This study paves the way for the use of a top-down proteomics strategy in barley for investigating more complex traits such as malting quality. |
