Evaluation of different protein extraction methods for banana (<i>Musa</i> spp.) root proteome analysis by two-dimensional electrophoresis

Abstract

Four protocols <i style="mso-bidi-font-style: normal">viz</i>., the trichloroacetic acid-acetone (TCA), phenol-ammonium acetate (PAA), phenol/SDS-ammonium acetate (PSA) and trisbase-acetone (TBA) were evaluated with modifications for protein extraction from banana (Grand Naine) roots, considered as recalcitrant tissues for proteomic analysis. The <span style="mso-bidi-font-weight:bold">two-dimensional electrophoresis<span style="mso-bidi-font-weight:bold"> (2-DE) separated proteins were compared based on protein yield, number of resolved proteins, sum of spot quantity, average spot intensity and proteins resolved in 4-7 p<i style="mso-bidi-font-style:normal">I</i> range. The PAA protocol yielded more proteins (0.89 mg/g of tissues) and protein spots (584) in 2-DE gel than TCA and other protocols. Also, the PAA protocol was superior in terms of sum of total spot quantity and average spot intensity than TCA and other protocols, suggesting phenol as extractant and ammonium acetate as precipitant of proteins were the most suitable for banana rooteomics analysis by 2-DE. In addition, 1:3 ratios of root tissue to extraction buffer and overnight protein precipitation were most efficient to obtain maximum protein yield.

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