Daniel J. Cosgrove —BIO-PROTOCOL

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Tian Zhang Bâtiment 2, INRA, Institute Jean-Pierre Bourgin, France, France,
2 protocols

Yong Bum Park Department of Biology and Center for LignoCellulose Structure and Formation, 208 Mueller Laboratory, Pennsylvania State University, USA, USA,
1 protocol

Daniel Durachko Department of Biology and Center for LignoCellulose Structure and Formation, 208 Mueller Laboratory, Pennsylvania State University, USA, USA,
1 protocol

Daniel J. Cosgrove

Department of Biology and Center for LignoCellulose Structure and Formation, 208 Mueller Laboratory, Pennsylvania State University, USA, USA,

2 Author merit
Reviewer merit
Editor merit

Education

PhD, Stanford University, USA

Current position

Professor, Biology, Penn State University, USA

Publications

https://scholar.google.com/citations?user=85KkItgAAAAJ&hl=en

2 Protocols published

Biomechanical Characterization of Onion Epidermal Cell Walls

Authors: Daniel M. Durachko, Yong Bum Park, Tian Zhang and Daniel J. Cosgrove, date: 12/20/2017, view: 5380, Q&A: 1

Here we describe two experimental protocols to measure the biomechanical properties of primary (growing) plant cell walls, with a focus on analyzing cell wall epidermal strips of onion scales. The first protocol measures cell wall creep ...

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William Nicolas

California Institute of Technology

Dear authors, I am currently trying to produce onion epidermal cell wall peels, exactly like in your Nature plants paper. This brought me to your detailed Bio-protocol paper. First of all, The pH at which the HEPES solution must be is not clear at all. In the materials in reagents section, it is said to be pH4.5, then in the recipe title, it is again 4.5 but in the description you say bring it to 6.8. Then figure 4, you say the peels are floating in HEPES buffer pH6.8. Moreover in the associated Nature plants paper, you say make a HEPES buffer solution pH7 + Tween-20 0.1%, which is not mentionned in this present protocol. I am especially asking this question because my first trials went quite well during the peeling stage, but when layed in the buffer, the peels would inevitably curl up into little tubes really hard to lay flat on a microscopy slide and the fact that this behaviour is not mentionned in your protocol leads me to think that you have not encountered such problem. I am thus suspecting my buffer, which does not contain the Tween-20 and is at pH6.8. In brief can you clear out the HEPES and sodium acetate buffer recipes please? Cheers! William Nicolas

1/24/2018 9:48:51 AM Reply

Ok thanks a lot, putting them with the cuticle facing up fixed the curling problem. Staining with DAPI also seems to indeed show the absence of the nuclei in the peeled part. The nuclei nonetheless remain in the thicker bordering parts, which is good. Thanks for the advice ! William Nicolas

2018-01-25 17:29:31

Daniel Cosgrove

Department of Biology and Center for LignoCellulose Structure and Formation, 208 Mueller Laboratory, Pennsylvania State University, USA, USA,

Thanks for your query and we are happy to clarify. In the Materials and Recipes sections, the pH of the HEPES and acetate buffers were incorrectly amended during the last stage of bio-protocol publication, and we missed this editorial change. HEPES buffer should be pH 6.8 and acetate buffer should be pH 4.5, throughout. Sorry for the confusion and we will ask bio-protocol to correct these typos. About curling: indeed the peeled strips have a strong tendency to curl. As described in step A.8 and shown in Figure 4B, it is important to leave chunks of subepidermal tissue at the two ends of the peel (like barbells). This helps a lot with the handling. Also, when you float the peels on a solution, ALWAYS place them with cuticle side up (facing the air). If you place the cuticle side down, they will curl. The power of surface tension of water! With a fine pair of forceps they can be uncurled under a dissecting scope, but it is challenging. The issue with curling should have been made clearer in the protocol. The curling behavior is not likely related to the solution composition. Also, we note that the tendency to curl varies greatly among onions, so try more than one onion. Good luck.

2018-01-25 08:05:56

Preparation of Onion Epidermal Cell Walls for Imaging by Atomic Force Microscopy (AFM)

Authors: Tian Zhang and Daniel J. Cosgrove, date: 12/20/2017, view: 6114, Q&A: 0

The growing plant cell wall is comprised of long, thin cellulose microfibrils embedded in a hydrated matrix of polysaccharides and glycoproteins. These components are typically constructed in layers (lamellae) on the inner surface of the cell wall, ...

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