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Evaluation of Three Protein-Extraction Methods for Proteome Analysis of Maize Leaf Midrib, a Compound Tissue Rich in Sclerenchyma Cells

Leaf morphology is closely related to the growth and development of maize (Zea mays L.) plants and final kernel production. As an important part of the maize leaf, the midrib holds leaf blades in the aerial position for maximum sunlight capture. Leaf midribs of adult plants contain substantial scler...

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Autores principales: Wang, Ning, Wu, Xiaolin, Ku, Lixia, Chen, Yanhui, Wang, Wei
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2016
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4905967/
https://www.ncbi.nlm.nih.gov/pubmed/27379139
http://dx.doi.org/10.3389/fpls.2016.00856
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author Wang, Ning
Wu, Xiaolin
Ku, Lixia
Chen, Yanhui
Wang, Wei
author_facet Wang, Ning
Wu, Xiaolin
Ku, Lixia
Chen, Yanhui
Wang, Wei
author_sort Wang, Ning
collection PubMed
description Leaf morphology is closely related to the growth and development of maize (Zea mays L.) plants and final kernel production. As an important part of the maize leaf, the midrib holds leaf blades in the aerial position for maximum sunlight capture. Leaf midribs of adult plants contain substantial sclerenchyma cells with heavily thickened and lignified secondary walls and have a high amount of phenolics, making protein extraction and proteome analysis difficult in leaf midrib tissue. In the present study, three protein-extraction methods that are commonly used in plant proteomics, i.e., phenol extraction, TCA/acetone extraction, and TCA/acetone/phenol extraction, were qualitatively and quantitatively evaluated based on 2DE maps and MS/MS analysis using the midribs of the 10th newly expanded leaves of maize plants. Microscopy revealed the existence of substantial amounts of sclerenchyma underneath maize midrib epidermises (particularly abaxial epidermises). The spot-number order obtained via 2DE mapping was as follows: phenol extraction (655) > TCA/acetone extraction (589) > TCA/acetone/phenol extraction (545). MS/MS analysis identified a total of 17 spots that exhibited 2-fold changes in abundance among the three methods (using phenol extraction as a control). Sixteen of the proteins identified were hydrophilic, with GRAVY values ranging from -0.026 to -0.487. For all three methods, we were able to obtain high-quality protein samples and good 2DE maps for the maize leaf midrib. However, phenol extraction produced a better 2DE map with greater resolution between spots, and TCA/acetone extraction produced higher protein yields. Thus, this paper includes a discussion regarding the possible reasons for differential protein extraction among the three methods. This study provides useful information that can be used to select suitable protein extraction methods for the proteome analysis of recalcitrant plant tissues that are rich in sclerenchyma cells.
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spelling pubmed-49059672016-07-04 Evaluation of Three Protein-Extraction Methods for Proteome Analysis of Maize Leaf Midrib, a Compound Tissue Rich in Sclerenchyma Cells Wang, Ning Wu, Xiaolin Ku, Lixia Chen, Yanhui Wang, Wei Front Plant Sci Plant Science Leaf morphology is closely related to the growth and development of maize (Zea mays L.) plants and final kernel production. As an important part of the maize leaf, the midrib holds leaf blades in the aerial position for maximum sunlight capture. Leaf midribs of adult plants contain substantial sclerenchyma cells with heavily thickened and lignified secondary walls and have a high amount of phenolics, making protein extraction and proteome analysis difficult in leaf midrib tissue. In the present study, three protein-extraction methods that are commonly used in plant proteomics, i.e., phenol extraction, TCA/acetone extraction, and TCA/acetone/phenol extraction, were qualitatively and quantitatively evaluated based on 2DE maps and MS/MS analysis using the midribs of the 10th newly expanded leaves of maize plants. Microscopy revealed the existence of substantial amounts of sclerenchyma underneath maize midrib epidermises (particularly abaxial epidermises). The spot-number order obtained via 2DE mapping was as follows: phenol extraction (655) > TCA/acetone extraction (589) > TCA/acetone/phenol extraction (545). MS/MS analysis identified a total of 17 spots that exhibited 2-fold changes in abundance among the three methods (using phenol extraction as a control). Sixteen of the proteins identified were hydrophilic, with GRAVY values ranging from -0.026 to -0.487. For all three methods, we were able to obtain high-quality protein samples and good 2DE maps for the maize leaf midrib. However, phenol extraction produced a better 2DE map with greater resolution between spots, and TCA/acetone extraction produced higher protein yields. Thus, this paper includes a discussion regarding the possible reasons for differential protein extraction among the three methods. This study provides useful information that can be used to select suitable protein extraction methods for the proteome analysis of recalcitrant plant tissues that are rich in sclerenchyma cells. Frontiers Media S.A. 2016-06-14 /pmc/articles/PMC4905967/ /pubmed/27379139 http://dx.doi.org/10.3389/fpls.2016.00856 Text en Copyright © 2016 Wang, Wu, Ku, Chen and Wang. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Plant Science
Wang, Ning
Wu, Xiaolin
Ku, Lixia
Chen, Yanhui
Wang, Wei
Evaluation of Three Protein-Extraction Methods for Proteome Analysis of Maize Leaf Midrib, a Compound Tissue Rich in Sclerenchyma Cells
title Evaluation of Three Protein-Extraction Methods for Proteome Analysis of Maize Leaf Midrib, a Compound Tissue Rich in Sclerenchyma Cells
title_full Evaluation of Three Protein-Extraction Methods for Proteome Analysis of Maize Leaf Midrib, a Compound Tissue Rich in Sclerenchyma Cells
title_fullStr Evaluation of Three Protein-Extraction Methods for Proteome Analysis of Maize Leaf Midrib, a Compound Tissue Rich in Sclerenchyma Cells
title_full_unstemmed Evaluation of Three Protein-Extraction Methods for Proteome Analysis of Maize Leaf Midrib, a Compound Tissue Rich in Sclerenchyma Cells
title_short Evaluation of Three Protein-Extraction Methods for Proteome Analysis of Maize Leaf Midrib, a Compound Tissue Rich in Sclerenchyma Cells
title_sort evaluation of three protein-extraction methods for proteome analysis of maize leaf midrib, a compound tissue rich in sclerenchyma cells
topic Plant Science
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4905967/
https://www.ncbi.nlm.nih.gov/pubmed/27379139
http://dx.doi.org/10.3389/fpls.2016.00856
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