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Proteomic analysis of cotyledonary explants during shoot organogenesis in Vigna radiata

Abstract

Vigna radiata or mungbean belongs to the legume family of plants. Mature mungbean seeds are rich source of dietary proteins for human nutrition. The present study was aimed to analyze the comparative protein profiles of two cotyledon types, Cot and Cot E, prior to and during early time points of shoot morphogenic induction to understand the unique differential regeneration response in these two explant types which was reported earlier. These explants were grown separately in vitro on the shoot induction medium (SIM) containing Gamborg’s B5 basal nutrient composition supplemented with 15 μM N6-benzyladenine. Isolation and characterization of the proteomes from Cot and Cot E explants at different time points, during early events of shoot differentiation, were performed using two dimensional gel electrophoresis following matrix assisted laser desorption-ionization tandem mass spectrometry. A total of 112 differentially identified proteins were classified according to their putative biological function. The differential control of protein synthesis between these explants under control condition, i.e. before in vitro culture, was also noted. In Cot E explants SIM induced prompt acquisition of competence for direct shoot morphogenesis probably through fast phytohormone signaling. Over accumulated proteins in Cot E indicated stimulation of several metabolic and associated pathways earlier than Cot explants. Abundance of stress and defense related proteins in Cot E explants was presumably to cope up with stressful cultural condition. Enhanced accumulation of folding-assisted proteins involved in organogenesis mediated cellular reprogramming in Cot E explants contributed further in rapid and efficient regeneration responsiveness.

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Abbreviations

SIM:

Shoot induction medium

2-D GE:

Two-dimensional gel electrophoresis

BA:

N6-benzyladenine

BSA:

Bovine serum albumin

Cv:

Cultivar

CV:

Coefficient of variation

DTT:

Dithiothreitol

IEF:

Isoelectric focusing

MALDI TOF MS:

Matrix assisted laser desorption-ionization time-of-flight mass spectrometry

MEV:

MultiExperiment viewer

MOWSE:

Molecular weight search

PMSF:

Phenylmethylsulfonyl fluoride

RT-PCR:

Reverse transcription polymerase chain reaction

SDS PAGE:

Sodium dodecyl sulphate polyacrylamide gel electrophoresis

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Acknowledgments

Authors are thankful to the Department of Science and Technology, Government of India (DST Sanction no. SR/SO/PS-58/05) for constant financial support in this area of research; and to the Director, Bose Institute for providing all infrastructural facilities and a Senior Research Fellowship to SG. The proteomic facilities provided by DST through IRHPA project (IR/SO/LF02/2002) are thankfully acknowledged.

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Correspondence to Amita Pal.

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11240_2013_340_MOESM1_ESM.jpg

Supplementary Fig. 1. Involvement of the identified proteins in the Glycolysis/Gluconeogenesis pathway assigned by KEGG database in Blast2go software. (JPEG 417 kb)

11240_2013_340_MOESM2_ESM.jpg

Supplementary Fig. 2. Involvement of the identified protein in the tricarboxylic acid cycle pathway assigned by KEGG database in Blast2go software. (JPEG 354 kb)

Supplementary material 3 (DOC 199 kb)

Supplementary material 4 (DOC 34 kb)

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Ghosh, S., Pal, A. Proteomic analysis of cotyledonary explants during shoot organogenesis in Vigna radiata . Plant Cell Tiss Organ Cult 115, 55–68 (2013). https://doi.org/10.1007/s11240-013-0340-0

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Keywords

  • Cytokinin
  • MALDI-TOF-TOF
  • Proteomics
  • Shoot regeneration
  • Vigna radiata