Этикетка без На месте Изображений одревеснения в стенах растительной клетки
Summary
Метод, основанный на конфокальной микроскопии комбинационного представлена, что позволяет без наклеек визуализации лигнина в стенках растительной клетки и сравнение одревеснения в различных тканей, образцов или видов.
Abstract
Meeting growing energy demands safely and efficiently is a pressing global challenge. Therefore, research into biofuels production that seeks to find cost-effective and sustainable solutions has become a topical and critical task. Lignocellulosic biomass is poised to become the primary source of biomass for the conversion to liquid biofuels1-6. However, the recalcitrance of these plant cell wall materials to cost-effective and efficient degradation presents a major impediment for their use in the production of biofuels and chemicals4. In particular, lignin, a complex and irregular poly-phenylpropanoid heteropolymer, becomes problematic to the postharvest deconstruction of lignocellulosic biomass. For example in biomass conversion for biofuels, it inhibits saccharification in processes aimed at producing simple sugars for fermentation7. The effective use of plant biomass for industrial purposes is in fact largely dependent on the extent to which the plant cell wall is lignified. The removal of lignin is a costly and limiting factor8 and lignin has therefore become a key plant breeding and genetic engineering target in order to improve cell wall conversion.
Analytical tools that permit the accurate rapid characterization of lignification of plant cell walls become increasingly important for evaluating a large number of breeding populations. Extractive procedures for the isolation of native components such as lignin are inevitably destructive, bringing about significant chemical and structural modifications9-11. Analytical chemical in situ methods are thus invaluable tools for the compositional and structural characterization of lignocellulosic materials. Raman microscopy is a technique that relies on inelastic or Raman scattering of monochromatic light, like that from a laser, where the shift in energy of the laser photons is related to molecular vibrations and presents an intrinsic label-free molecular “fingerprint” of the sample. Raman microscopy can afford non-destructive and comparatively inexpensive measurements with minimal sample preparation, giving insights into chemical composition and molecular structure in a close to native state. Chemical imaging by confocal Raman microscopy has been previously used for the visualization of the spatial distribution of cellulose and lignin in wood cell walls12-14. Based on these earlier results, we have recently adopted this method to compare lignification in wild type and lignin-deficient transgenic Populus trichocarpa (black cottonwood) stem wood15. Analyzing the lignin Raman bands16,17 in the spectral region between 1,600 and 1,700 cm-1, lignin signal intensity and localization were mapped in situ. Our approach visualized differences in lignin content, localization, and chemical composition. Most recently, we demonstrated Raman imaging of cell wall polymers in Arabidopsis thaliana with lateral resolution that is sub-μm18. Here, this method is presented affording visualization of lignin in plant cell walls and comparison of lignification in different tissues, samples or species without staining or labeling of the tissues.
Protocol
Discussion
Лигноцеллюлозной материалы являются иерархическими и гетерогенных как с точки зрения структуры и состава. Для углубленного характеристика аналитических инструментов, которые имеют химическую чувствительность, пространственное разрешение, и что дают представление этих материалов ?…
Divulgazioni
The authors have nothing to disclose.
Acknowledgements
Мы благодарим Эндрю Кэрролл, светлая Chaibang, Purbasha Саркар (Energy Biosciences институт, Беркли), Бахрам Парвин (Lawrence Berkeley National Laboratory) и Винсент Л. Чан (North Carolina State University) за плодотворное сотрудничество и полезные обсуждения. Эта работа была поддержана Энергия биологических наук института. Работа в молекулярной Литейное при поддержке Управления науки, Управление основной энергии наук, из Министерства энергетики США по контракту № DE-AC02-05CH1123.
Materials
| Material Name | Tipo | Company | Catalogue Number | Comment |
|---|---|---|---|---|
| microscope slides | ||||
| cover slips | ||||
| D2O | ||||
| nail polish | ||||
| immersion oil | ||||
| tweezers | ||||
| pointed brush | ||||
| microtome | ||||
| confocal Raman microscope |
Riferimenti
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