Isolation of Whole Esophageal Gland Cells from Plant-Parasitic Nematodes for Transcriptome Analyses and Effector Identification
Open Access
- 1 January 2013
- journal article
- research article
- Published by Scientific Societies in Molecular Plant-Microbe Interactions®
- Vol. 26 (1), 31-35
- https://doi.org/10.1094/mpmi-05-12-0121-fi
Abstract
Esophageal glands of plant-parasitic nematodes are highly specialized cells whose gene expression products include secreted effector proteins, which govern nematode parasitism of host plants. Therefore, elucidating the transcriptomes of esophageal glands with the goal of identifying nematode effectors is a promising avenue to understanding nematode parasitism and its evolutionary origins as well as to devising nematode control strategies. We have developed a method to separate and isolate individual esophageal gland cells from multiple species of plant-parasitic nematodes while preserving RNA quality. We have used such isolated gland cells for transcriptome analysis via high-throughput DNA sequencing. This method relies on the differential histochemical staining of the gland cells after homogenization of phytonematode tissues. Total RNA was extracted from whole gland cells isolated from eight different plant-parasitic nematode species. To validate this approach, the isolated RNA from three plant-parasitic nematode species—Globodera rostochiensis, Pratylenchus penetrans, and Radopholus similis—was amplified, gel purified, and used for 454 sequencing. We obtained 456,801 total reads with an average read length of 409 bp. Sequence analyses revealed the presence of homologs of previously known nematode effectors in these libraries, thus validating our approach. These data provide compelling evidence that this technical advance can be used to relatively easily and expediently discover effector repertoires of plant-parasitic nematodes.Keywords
This publication has 12 references indexed in Scilit:
- A root‐knot nematode‐secreted protein is injected into giant cells and targeted to the nucleiNew Phytologist, 2012
- Multiple lateral gene transfers and duplications have promoted plant parasitism ability in nematodesProceedings of the National Academy of Sciences of the United States of America, 2010
- Methods for extraction, processing and detection of plant and soil nematodes.Published by CABI Publishing ,2005
- Getting to the roots of parasitism by nematodesTrends in Parasitology, 2004
- The Parasitome of the Phytonematode Heterodera glycinesMolecular Plant-Microbe Interactions®, 2003
- A Profile of Putative Parasitism Genes Expressed in the Esophageal Gland Cells of the Root-knot Nematode Meloidogyne incognitaMolecular Plant-Microbe Interactions®, 2003
- Secrets in secretions: genes that control nematode parasitism of plantsBrazilian Journal of Plant Physiology, 2002
- Single pass cDNA sequencing - a powerful tool to analyse gene expression in preparasitic juveniles of the southern root-knot nematode Meloidogyne incognitaNematology, 2001
- Ultrastructure of esophageal glands and their secretory granules in the root-knot nematodeMeloidogyne incognitaProtoplasma, 1990
- Disease-Inducing Secretions of Plant-Parasitic NematodesAnnual Review of Phytopathology, 1989