Histochemical Staining of <em>Arabidopsis thaliana</em> Secondary Cell Wall Elements

Abstract

Arabidopsis thaliana is a model organism commonly used to understand and manipulate various cellular processes in plants, and it has been used extensively in the study of secondary cell wall formation. Secondary cell wall deposition occurs after the primary cell wall is laid down, a process carried out exclusively by specialized cells such as those forming vessel and fiber tissues. Most secondary cell walls are composed of cellulose (40-50%), hemicellulose (25-30%), and lignin (20-30%). Several mutations affecting secondary cell wall biosynthesis have been isolated, and the corresponding mutants may or may not exhibit obvious biochemical composition changes or visual phenotypes since these mutations could be masked by compensatory responses. Staining procedures have historically been used to show differences on a cellular basis. These methods are exclusively visual means of analysis; nevertheless their role in rapid and critical analysis is of great importance. Congo red and calcofluor white are stains used to detect polysaccharides, whereas Mäule and phloroglucinol are commonly used to determine differences in lignin, and toluidine blue O is used to differentially stain polysaccharides and lignin. The seemingly simple techniques of sectioning, staining, and imaging can be a challenge for beginners. Starting with sample preparation using the A. thaliana model, this study details the protocols of a variety of staining methodologies that can be easily implemented for observation of cell and tissue organization in secondary cell walls of plants.

Elicitors and defense gene induction in plants with altered lignin compositions

“…S1). To check the cellular distribution of the lignin, we examined cross‐sections of inflorescence stems for lignin autofluorescence under UV light, and by Mäule staining, which yields a red coloration with S lignin (Pradhan Mitra & Loque, ) (Fig. S2).…”

“…Ten milligrams of AIRs extracted from stem tissue were subjected to lignin composition analysis using the thioacidolysis method as reported previously (Zhao et al ., ), and cellular localization of lignin was determined by Mäule staining of cross‐sections of inflorescence stems (Pradhan Mitra & Loque, ).…”

Cotton GhMYB7 is predominantly expressed in developing fibers and regulates secondary cell wall biosynthesis in transgenic Arabidopsis

“…Next, we examined if overexpression of GhMYB7 caused ectopic deposition of SCW cellulose in transgenic Arabidopsis. Calcofluor white, Congo red, and S4B were used to image the cellulose distribution in plant cell walls (An-derson et al, 2010;Pradhan et al, 2014). As shown in Figure 3M-R, histological staining of cellulose in stems of the transgenic lines revealed that ectopic deposition of cellulose occurred in GhMYB7 transgenic stem epidermal cells, whereas SCW thickening was not detected in stem epidermal cells of wild-type plants.…”

“…For lignin staining, sections were stained with either phloroglucinol HCl or Maule reagent as described previously (Pradhan et al, 2014). Positive results for phloroglucinol staining indicate the presence of guaiacyl-lignin, whereas those for Maule reagent staining indicate the presence of syringyl-lignin.…”

Composite plants for a composite plant: An efficient protocol for root studies in the sunflower using composite plants approach

“…Sunflower is described as normally susceptible to low temperatures and salinity, but with a relative tolerance to drought stress due to its highly explorative root system. However, available information on gene expression in response to abiotic stresses in sunflower is still limited to few works (Pradhan Mitra et al 2014; Vitha et al 1995)…”