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Summary

Abstract

Introduction

Protocol

Representative Results

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Acknowledgements

Materials

References

Environment

Quantifying Plant Soluble Protein and Digestible Carbohydrate Content, Using Corn (Zea mays) As an Exemplar

Published: August 6th, 2018

DOI:

10.3791/58164

1Department of Entomology, Texas A&M University, 2Department of Entomology, University of Minnesota, 3Department of Entomology, University of Kentucky

The protocols described herein provide a clear and approachable methodology for measuring soluble protein and digestible (non-structural) carbohydrate content in plant tissues. The ability to quantify these two plant macronutrients has significant implications for advancing the fields of plant physiology, nutritional ecology, plant-herbivore interactions and food-web ecology.

Elemental data are commonly used to infer plant quality as a resource to herbivores. However, the ubiquity of carbon in biomolecules, the presence of nitrogen-containing plant defensive compounds, and variation in species-specific correlations between nitrogen and plant protein content all limit the accuracy of these inferences. Additionally, research focused on plant and/or herbivore physiology require a level of accuracy that is not achieved using generalized correlations. The methods presented here offer researchers a clear and rapid protocol for directly measuring plant soluble proteins and digestible carbohydrates, the two plant macronutrients most closely tied to animal physiological performance. The protocols combine well characterized colorimetric assays with optimized plant-specific digestion steps to provide precise and reproducible results. Our analyses of different sweet corn tissues show that these assays have the sensitivity to detect variation in plant soluble protein and digestible carbohydrate content across multiple spatial scales. These include between-plant differences across growing regions and plant species or varieties, as well as within-plant differences in tissue type and even positional differences within the same tissue. Combining soluble protein and digestible carbohydrate content with elemental data also has the potential to provide new opportunities in plant biology to connect plant mineral nutrition with plant physiological processes. These analyses also help generate the soluble protein and digestible carbohydrate data needed to study nutritional ecology, plant-herbivore interactions and food-web dynamics, which will in turn enhance physiology and ecological research.

Plant biomass forms the foundation of virtually all terrestrial food-webs. Plants acquire nutritional elements from the soil through their roots systems and utilize sunlight in their foliar tissues to synthesize biomolecules. In particular, carbon and nitrogen are used to create carbohydrates, proteins (comprised of amino acids), and lipids that are needed to build plant biomass (it should be noted that in plant physiology the term "macronutrient" often refers to soil elements, such as N, P, K, and S, however, throughout this paper this term will refer to biomolecules, such as proteins, carbohydrates, and lipids). When herbivores consume plant material, the ma....

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1. Plant Collection and Processing

  1. Collect and process plant samples
    1. After collecting plant samples, flash-freeze samples by dipping plant material into liquid nitrogen with forceps and store at -80 °C. If the plant samples collected are too large to flash-freeze, quickly cool the samples using dry ice and transfer to a -80 °C freezer as soon as possible. The macronutrient content of plant material can change after tissues are separated from the plant, so it is important to freeze plant .......

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To show the usefulness of these methods, we analyzed the soluble protein and digestible carbohydrate content of four different field and sweetcorn tissues that serve as distinct potential nutritional resources for insect herbivores. We collected ears of corn from three agricultural regions in the United States (Minnesota, North Carolina, and Texas), encompassing five different varieties of sweet corn (i.e., genotypes) and one variety of field corn as an outgroup. Table 3<.......

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By combining well-established colorimetric assays with effective plant-specific extraction protocols, the assays demonstrated here provide a reasonable and accurate method for measuring plant soluble protein and digestible carbohydrate content. Our results using corn as an exemplar illustrates how these protocols can be used to obtain precise measurements across different biologically-relevant spatial scales. For example, we were able to detect differences in plant soluble protein and digestible carbohydrate content betw.......

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Thanks to all of our collaborators who have assisted with sweet corn field collections, including Dominic Reisig and Dan Mott at North Carolina State University, and Pat Porter at Texas A& M University in Lubbock, TX. Thanks to Fiona Clissold for helping to optimize the protocols and for providing edits to this manuscript. This work was supported in part by the Texas A& M C. Everette Salyer Fellowship (Department of Entomology) and the Biotechnology Risk Assessment Grant Program competitive grant no. 2015-33522-24099 from the U.S. Department of Agriculture (awarded to GAS and STB).

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Name Company Catalog Number Comments
microplate reader (spectrophotometer) Bio-Rad Model 680 XR
Bio-Rad Protein Assay Dye Reagent concentrate Bio-Rad #5000006 450mL

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