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Article
Flow Cytometric Analysis and Sorting of Heterodera glycines Eggs
Journal of Nematology
  • G. L. Tylka, Iowa State University
  • T. L. Niblack, University of Missouri
  • T. C. Walk, University of Missouri
  • K. R. Harkins, Iowa State University
  • L. Barnett, University of Missouri
  • N. K. Baker, Iowa State University
Document Type
Article
Publication Version
Published Version
Publication Date
12-1-1993
Abstract

A nondestructive technique was developed to characterize and separate eggs of soybean cyst nematode, Heterodera glycines, by developmental stage using flow cytometry. Eggs from cysts cultured on susceptible soybean roots were suspended in 0.1% xanthan gum or 59% sucrose and loaded into either a Coulter EPICS 752 or EPICS 753 flow cytometer. Eggs were analyzed and sorted according to forward angle and 90º light scatter, flow cytometric parameters that are relative measures of object size and granularity, respectively. Mature eggs containing vermiform juveniles were less granular and slightly larger than eggs in earlier stages of embryogeny, allowing for separation of mature eggs from immature eggs. The effectiveness of flow cytometric sorting was evaluated by comparing the developmental stages of subpopulations of unsorted and sorted eggs. Of a subpopulation of unsorted eggs, 62% contained vermiform juveniles, whereas 85 to 95% of sorted subpopulations of larger, less granular eggs contained vermiform juveniles. Suspending H. glycines eggs in 0.1% xanthan gum or 59% sucrose for flow cytometric analysis had no effect on subsequent egg hatch in vitro. This technique is an efficient and effective means to collect large, relatively homogeneous quantities of H. glycines eggs in early or late embryogeny, and would likely be useful for analyzing and sorting eggs of other nematode species for use in developmental, genetic, or physiological research, or for identification and collection of parasitized eggs. Key words: egg development, embryogeny, flow cytometry, Heterodera glycines, method, nematode, nematode egg sorting, soybean cyst nematode.

Comments

This article is published as Tylka, G. L., T. L. Niblack, T. C. Walk, K. R. Harkins, L. Barnett, and N. K. Baker. "Flow Cytometric Analysis and Sorting of Heterodera glycines Eggs." Journal of Nematology 25, no. 4: 596-602. Posted with permission.

Copyright Owner
The Society of Nematologists
Language
en
File Format
application/pdf
Citation Information
G. L. Tylka, T. L. Niblack, T. C. Walk, K. R. Harkins, et al.. "Flow Cytometric Analysis and Sorting of Heterodera glycines Eggs" Journal of Nematology Vol. 25 Iss. 4 (1993) p. 596 - 602
Available at: http://works.bepress.com/gregory-tylka/201/