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<article dtd-version="3.0" article-type="Invited Methods Article - JoVE Produced Video">
  <front>
    <journal-meta>
      <journal-id journal-id-type="publisher">jove</journal-id>
      <journal-id journal-id-type="email">editorial-office@jove.com</journal-id>
      <journal-title-group>
        <journal-title>Journal of Visualized Experiments</journal-title>
      </journal-title-group>
      <issn pub-type="ppub">1940-087X</issn>
      <issn pub-type="epub"></issn>
    </journal-meta>
    <article-meta>
      <article-id pub-id-type="publisher-id">JoVE56424</article-id>
      <article-id pub-id-type="manuscript">JoVE56424</article-id>
      <article-categories>
        <subj-group subj-group-type="Article Type">
          <subject>Invited Methods Article - JoVE Produced Video</subject>
        </subj-group>
        <subj-group subj-group-type="Category">
          <subject>JoVE Biochemistry</subject>
        </subj-group>
        <subj-group subj-group-type="Classification">
          <subject>Cell Wall</subject>
          <subject>Biopolymers</subject>
          <subject>Glycoside Hydrolases</subject>
          <subject>Glycosyltransferases</subject>
          <subject>Polysaccharides</subject>
          <subject>Electrophoresis</subject>
          <subject>Glycosylation</subject>
          <subject>Glycomics</subject>
        </subj-group>
      </article-categories>
      <title-group>
        <article-title>Structural analysis of mannan cell wall polysaccharides in plants using PACE.</article-title>
      </title-group>
      <contrib-group>
        <contrib contrib-type="author">
          <role content-type="1" />
          <name>
            <surname>Pidatala</surname>
            <given-names>Venkataramana  R</given-names>
          </name>
          <email>ramana@lbl.gov</email>
          <xref ref-type="aff" rid="aff1" />
        </contrib>
        <contrib contrib-type="author">
          <role content-type="2" />
          <name>
            <surname>Mahboubi</surname>
            <given-names>Amir</given-names>
          </name>
          <email>mmahboubi@lbl.gov</email>
          <xref ref-type="aff" rid="aff2" />
        </contrib>
        <contrib contrib-type="author" corresp="yes">
          <role content-type="3" />
          <name>
            <surname>Mortimer</surname>
            <given-names>Jenny  C</given-names>
            <prefix>Dr.</prefix>
          </name>
          <degrees>Ph.D.</degrees>
          <email>jcmortimer@lbl.gov</email>
          <xref ref-type="aff" rid="aff3" />
          <xref ref-type="aff" rid="orcid6" />
        </contrib>
        <contrib contrib-type="editor">
          <name>
            <surname>Kolski-Andreaco</surname>
            <given-names>Aaron</given-names>
          </name>
          <degrees>Ph.D.</degrees>
          <email>aaronk.andreaco@jove.com</email>
          <xref ref-type="aff" rid="aff4" />
        </contrib>
        <aff id="aff1">
          <institution>E O Lawrence Berkeley National Laboratory</institution>
          <addr-line content-type="addrline1">1 Cyclotron Road</addr-line>
          <country>UNITED STATES</country>
          <addr-line content-type="city">Berkeley</addr-line>
          <addr-line content-type="zipcode">94720</addr-line>
        </aff>
        <aff id="aff2">
          <institution>E O Lawrence Berkeley National Laboratory</institution>
          <addr-line content-type="addrline1">1 Cyclotron Road</addr-line>
          <country>UNITED STATES</country>
          <addr-line content-type="city">Berkeley</addr-line>
          <addr-line content-type="zipcode">CA</addr-line>
        </aff>
        <aff id="aff3">
          <institution>E O Lawrence Berkeley National Laboratory</institution>
          <institution content-type="position">Director of Plant Systems Biology, Joint BioEnergy Institute</institution>
          <addr-line content-type="addrline1">1 Cyclotron Road</addr-line>
          <country>UNITED STATES</country>
          <addr-line content-type="city">Berkeley</addr-line>
          <addr-line content-type="state">CA</addr-line>
          <addr-line content-type="zipcode">94720</addr-line>
          <phone content-type="primary">+1-510-486-6627</phone>
          <institution>E O Lawrence Berkeley National Laboratory</institution>
        </aff>
        <aff id="orcid6">0000-0001-6624-636X</aff>
        <aff id="aff4">
          <institution>JoVE</institution>
          <addr-line>1 Alewife Center</addr-line>
          <addr-line>Suite 200</addr-line>
          <addr-line>Cambridge, MA  02140</addr-line>
          <country>UNITED STATES</country>
        </aff>
      </contrib-group>
      <history>
        <date date-type="received">
          <day>13</day>
          <month>04</month>
          <year>2017</year>
        </date>
        <date date-type="rev-recd">
          <day>12</day>
          <month>06</month>
          <year>2017</year>
        </date>
        <date date-type="accepted">
          <day>20</day>
          <month>06</month>
          <year>2017</year>
        </date>
      </history>
      <abstract>
        <p>Plant cell wall polysaccharides are notoriously difficult to analyze, and most methods require expensive equipment, skilled operators, and large amounts of purified material. Here, we describe a simple method for gaining detailed polysaccharide structural information, including resolution of structural isomers. For polysaccharide analysis by gel electrophoresis (PACE), plant cell wall material is hydrolyzed with glycosyl hydrolases specific to the polysaccharide of interest (e.g. mannanases for mannan). Large format polyacrylamide gels are then used to separate the released oligosaccharides, which have been fluorescently labelled. Gels can be visualized with a modified "geldoc" system. The resulting oligosaccharide fingerprint can either be compared qualitatively or, with replication, quantitatively. Linkage and branching information can be established using additional glycosyl hydrolases (e.g. mannosidases and galactosidases). Whilst this protocol describes a method for analyzing glucomannan structure, it can be applied to any polysaccharide for which characterized glycosyl hydrolases exist. Alternatively, it can be used to characterize novel glycosyl hydrolases using defined polysaccharide substrates.</p>
      </abstract>
      <kwd-group>
        <kwd>Gel electrophoresis</kwd>
        <kwd>polysaccharides</kwd>
        <kwd>glycosyl hydrolases</kwd>
        <kwd>mannanases</kwd>
        <kwd>mannan</kwd>
        <kwd>glucomannan</kwd>
        <kwd>cell wall</kwd>
        <kwd>hemicellulose</kwd>
        <kwd>biofuels.</kwd>
      </kwd-group>
      <funding-group />
      <counts>
        <fig-count count="0" />
      </counts>
      <custom-meta-group>
        <custom-meta>
          <meta-name>Please indicate whether this article will be standard or open access. </meta-name>
          <meta-value>Open Access (US$4,200);</meta-value>
        </custom-meta>
        <custom-meta>
          <meta-name>Please indicate the &lt;b&gt; full address&lt;/b&gt; at which this article will be &lt;b&gt;filmed&lt;/b&gt;.</meta-name>
          <meta-value>5885 Hollis St., 4th Floor, Emeryville, CA 94608</meta-value>
        </custom-meta>
        <custom-meta>
          <meta-name>If the invoicing address and point of contact are different than the corresponding author and filming address, please note the differences below.</meta-name>
          <meta-value>Mailing address:
1 Cyclotron Rd.
MS: 978-4121
Berkeley, CA 94720</meta-value>
        </custom-meta>
        <custom-meta>
          <meta-name>Is all of the written text in your manuscript original (neither plagiarized nor self-plagiarized)? If using previously published figures, are re-use permissions from the previous publishers obtained and cited in the figure legends?</meta-name>
          <meta-value>Yes, it's all original</meta-value>
        </custom-meta>
        <custom-meta>
          <meta-name>If this article is currently under review by another publication, please explain where.</meta-name>
          <meta-value>No</meta-value>
        </custom-meta>
        <custom-meta>
          <meta-name>Black and White Image Count</meta-name>
          <meta-value>0</meta-value>
        </custom-meta>
        <custom-meta>
          <meta-name>Color Image Count</meta-name>
          <meta-value>0</meta-value>
        </custom-meta>
        <custom-meta>
          <meta-name>Acquisitions Editor</meta-name>
          <meta-value>Benjamin Werth</meta-value>
        </custom-meta>
        <custom-meta>
          <meta-name>Assistant who entered lead</meta-name>
          <meta-value>Benjamin Werth</meta-value>
        </custom-meta>
      </custom-meta-group>
    </article-meta>
  </front>
  <body>
    <fig>
      <label>Figure 1</label>
      <graphic xlink:href="Fig1_Derivatization.pdf" xmlns:xlink="http://www.w3.org/1999/xlink" />
    </fig>
    <fig>
      <label>Figure 2</label>
      <graphic xlink:href="Fig2_cellwall.pdf" xmlns:xlink="http://www.w3.org/1999/xlink" />
    </fig>
    <table-wrap>
      <label>Table of Materials/Equipment</label>
      <graphic xlink:href="JoVE Materials List _Mortimer.xlsx" xmlns:xlink="http://www.w3.org/1999/xlink" />
    </table-wrap>
    <fig>
      <label>Figure 2</label>
      <graphic xlink:href="Fig2_cellwall_rev.pdf" xmlns:xlink="http://www.w3.org/1999/xlink" />
    </fig>
    <table-wrap>
      <label>Table of Materials/Equipment</label>
      <graphic xlink:href="JoVE Materials List revised.xlsx" xmlns:xlink="http://www.w3.org/1999/xlink" />
    </table-wrap>
  </body>
</article>