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Molecular Biotechnology

, Volume 22, Issue 1, pp 19–23 | Cite as

A gel diffusion assay for visualization and quantification of chitinase activity

  • Xiaohong Zou
  • Hiroyuki Nonogaki
  • Gregory E. Welbaum
Research

Abstract

Higher plants, bacteria, fungi, insects, and crustaceans all produce chitinases. Chitinase genes in many organisms are currently under investigation. Chitinase activity is usually assayed with radiolabeled or fluorogenic substrates. We developed a simple, inexpensive, nonradioactive gel-diffusion assay for chitinase that can be used to screen large numbers of samples. In this assay, chitinase diffuses from a small circular well cut in an agarose or agar gel containing the substrate glycol chitin, a soluble, modified form of chitin. Chitinase catalyzes the cleavage of glycol chitin as it diffuses through the gel, leaving a dark, unstained circular zone around the well, because the fluorescent dye calcofluor binds only to undigested chitin. Sample activities can be determined from linear regression of logstandard enzyme concentration versus the zone diameter of internal standards on each Petri dish used for a diffusion assay.

Index Entries

Chitinase gel diffusion assay enzyme activity 

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Copyright information

© Humana Press Inc 2002

Authors and Affiliations

  • Xiaohong Zou
    • 1
  • Hiroyuki Nonogaki
    • 2
  • Gregory E. Welbaum
    • 1
  1. 1.Department of Horticulture, Saunders HallVirginia TechBlacksburg
  2. 2.Department of Crop and Soil Sci.Oregon State UniversityCorvallis

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