An efficient protein extraction method for proteomic analysis of black pepper (Piper nigrum L.) and generation of protein map using nano LC-LTQ Orbitrap mass spectrometry

Abstract

Information on black pepper proteins can accelerate the basic understanding of crop response to various biotic, abiotic stresses as well as the crop improvement programmes. Here, we present the protocols for total leaf protein extraction from this non model, recalcitrant plant species P. nigrum after evaluating 5 different standard methods. The efficiency and suitability of the methods were evaluated using important parameters such as protein quantity and quality, solubility in IEF compatible buffer and streak free results, number of spots in 2DE gel. Modified lysis buffer and phenol method were found to be applicable methods for protein extraction from black pepper. Among two applicable methods, the phenol protocol is time-consuming, demanding the toxic phenol. Hence, we propose the modified lysis buffer method as superior, rapid method with no polyphenol contamination for total leaf protein extraction. This is a very first report which evaluated different methods and optimized the standards for the analysis of black pepper leaf proteome. Successful identification of randomly selected spots by nano LC-LTQ Orbitrap proves its compatibility to high throughput MS analysis which leads to the generation of leaf protein map, the first of its kind.