1. Volume 4, Issue 6, Pages 1024-1037 (November 2011)
The Cooperative Activities of CSLD2, CSLD3, and CSLD5 Are Required for Normal Arabidopsis Development 
Yin Lan , Verhertbruggen Yves , Oikawa Ai , Manisseri Chithra , Knierim Bernhard , Prak Lina , Jensen Jacob Krüger , Knox J. Paul , Auer Manfred , Willats William G.T. , Scheller Henrik Vibe  
Molecular Plant 
Volume 4, Issue 6, Pages (November 2011)
DOI: /mp/ssr026
Copyright © 2011 The Authors. All rights reserved. Terms and Conditions

2. Figure 1
The csld2/csld5, csld3/csld5 and csld2/csld3/csld5 Mutants All Displayed a Severely Restricted Growth Phenotype Compared to Col-0.
(A) 23-day-old plants.
(B) 5-day-old seedlings. At this magnification, root hairs can be observed in Col-0 and csld2/csld5 but not in csld3/csld5 and csld2/csld3/csld5.
Molecular Plant 2011 4, DOI: ( /mp/ssr026)
Copyright © 2011 The Authors. All rights reserved. Terms and Conditions

3. Figure 2 Tissue Analysis of Stems.
(A–F) Transverse stem sections stained with toluidine blue.
(G–L) Transmission electron microscopy micrographs showing xylem cells and cortical parenchyma cells (M). ‘TM’ designates the csld2/csld3/csld5 triple mutant. ep, epidermis; cp, cortical parenchyma; ed, endodermis; pf, phloem fibers; ph, phloem; if, interfascicular fibers; xy, xylem; pp, pith parenchyma. Black dashed, white dashed, and full black lines show the cortical parenchyma, the phloem, and the xylem, respectively. Arrowheads highlight the staining of toluidine blue of the secondary cell walls. The arrows in (M) show unknown bodies that only occur in the cortical parenchyma of the triple mutant. Scale bars: (A–D, F) = 30 μm; (E) = 40 μm; (G–L) = 4 μm; (P) = 5 μm.
Molecular Plant 2011 4, DOI: ( /mp/ssr026)
Copyright © 2011 The Authors. All rights reserved. Terms and Conditions

4. Figure 3
Vascular Patterning and Development in csld2/csld3/csld5 Are Abnormal.
(A, B) Cotyledons 8 and 16 d after germination for Col-0 and mutant, respectively.
(C, D) Rosette leaves 14 and 25 d after germination for Col-0 and mutant, respectively. Leaves were cleared using chloral hydrate to show the vascular pattern.
Molecular Plant 2011 4, DOI: ( /mp/ssr026)
Copyright © 2011 The Authors. All rights reserved. Terms and Conditions

5. Figure 4
Complementation Experiments with YFP–CSLD2, YFP–CSLD3, and YFP–CSLD5 Driven by 35S Promoter.
All roots imaged here were from 3-day-old seedlings grown on MS medium at 22°C. The arrows indicate deformed root hairs with swelling at the hair base and/or tip rupture.
(A) csld3 mutant.
(B–D) csld3 mutant transformed with YFP–CSLD2, YFP–CSLD3, and YFP–CSLD5 as indicated on top.
(E) csld2/csld3 mutant.
(F–H) csld2/csld3 mutant transformed with YFP–CSLD2, YFP–CSLD3, and YFP–CSLD5 as indicated on top. The fluorescence images (B2 corresponding to B1, C2 corresponding to C1, etc.) are included to demonstrate that the seedlings used for analysis expressed the transgenes at similar levels.
Molecular Plant 2011 4, DOI: ( /mp/ssr026)
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6. Figure 5
Staining and Immunodetection of Cell Wall Polysaccharides at the Surface of Intact Root Hairs of 7-Day-Old Seedlings. Bright field shows a representative root hair of each genotype. Calcofluor stains β-glucans, including crystalline cellulose. The LM21 rat monoclonal antibody binds to mannan. Arrowheads indicate the tip of the root hair. Scale bars: bright field = 10 mm; Calcofluor, LM21 = 20 mm.
Molecular Plant 2011 4, DOI: ( /mp/ssr026)
Copyright © 2011 The Authors. All rights reserved. Terms and Conditions

7. Figure 6
Staining and Immunodetection of Cell Wall Polysaccharides in Transverse Sections of Inflorescence Stems of 20-Day-Old Col-0 (Col-0 20), 40-Day-Old Col-0 (Col-0 40), and 40-Day-Old csld2/csld3/csld5 (TM). Resin-embedded stem sections were stained with Calcofluor or immunolabeled with monoclonal antibodies to homogalacturonan (LM19 and LM20), xylan (LM10), and mannan (LM21). Inserts are higher magnifications of phloem cells labeled by LM20 and epidermal cell labeled by LM19 in Col-0 40, and of xylem cells labeled by LM10 and intracellular labeling by LM21 in the young wild-type stem section. Scale bars: large micrographs = 50 μm; inserts = 5 μm.
Molecular Plant 2011 4, DOI: ( /mp/ssr026)
Copyright © 2011 The Authors. All rights reserved. Terms and Conditions

8. Figure 7
Glycosyltransferase Assays with Arabidopsis CSLD5 Heterologously Expressed in N. benthamiana. Microsomes of N. benthamiana leaves transiently expressing YFP–CSLD5 were assayed for glycosyltransferase activity using endogenous acceptors (A, B, D) or monosaccharides as acceptors (C). Experiments were performed using duplicated samples and the data represent mean ± SD from two to four independent experiments.
(A) Enzyme reactions were performed for 2 h with UDP–14C-Glc (694 Bq), UDP–14C-Gal (555 Bq), UDP–14C-Xyl (278 Bq), or GDP–14C-Man (555 Bq). The products were precipitated in 70% (v/v) EtOH and analyzed by liquid scintillation counting.
(B) The in vitro assay products obtained with GDP–14C-Man as substrate were treated with various hydrolytic enzymes. After treatment, EtOH was added to 70% (v/v), and the radioactivity in supernatant and precipitate was determined.
(C) Enzyme reactions were performed for 3 h with GDP–14C-Man (555 Bq) in the presence of monosaccharide acceptor (glucose or mannose). The products were purified by anion exchange chromatography and scintillation counted.
(D) Enzyme reactions were performed for 2 h with GDP–14C-Man (555 Bq, 50 μM) and/or GDP–14C-Glc (555 Bq, 50 μM). The Glc*ManS product was labeled with GDP–14C-Glc in the presence of 50 μM nonradioactive GDP–Man, and the GlcMan*S product was labeled with GDP–14C-Man in the presence of 50 μM nonradioactive GDP–Glc. The products were precipitated in 70% (v/v) EtOH and analyzed by liquid scintillation counting.
Molecular Plant 2011 4, DOI: ( /mp/ssr026)
Copyright © 2011 The Authors. All rights reserved. Terms and Conditions