Quantitative Detection of Double-Stranded RNA-Mediated Gene Silencing of Parasitism Genes in Heterodera glycines

Thumbnail Image
Date
2007-06-01
Authors
Sukno, Serenella
McCuiston, Jamie
Wong, Mui-Yun
Wang, Xiaohong
Thon, Michael
Hussey, Richard
Davis, Eric
Major Professor
Advisor
Committee Member
Journal Title
Journal ISSN
Volume Title
Publisher
Authors
Person
Baum, Thomas
Distinguished Professor
Research Projects
Organizational Units
Organizational Unit
Plant Pathology and Microbiology
The Department of Plant Pathology and Microbiology and the Department of Entomology officially merged as of September 1, 2022. The new department is known as the Department of Plant Pathology, Entomology, and Microbiology (PPEM). The overall mission of the Department is to benefit society through research, teaching, and extension activities that improve pest management and prevent disease. Collectively, the Department consists of about 100 faculty, staff, and students who are engaged in research, teaching, and extension activities that are central to the mission of the College of Agriculture and Life Sciences. The Department possesses state-of-the-art research and teaching facilities in the Advanced Research and Teaching Building and in Science II. In addition, research and extension activities are performed off-campus at the Field Extension Education Laboratory, the Horticulture Station, the Agriculture Engineering/Agronomy Farm, and several Research and Demonstration Farms located around the state. Furthermore, the Department houses the Plant and Insect Diagnostic Clinic, the Iowa Soybean Research Center, the Insect Zoo, and BugGuide. Several USDA-ARS scientists are also affiliated with the Department.
Journal Issue
Is Version Of
Versions
Series
Abstract

The introduction of a double-stranded RNA (dsRNA) into an organism to induce sequence-specific RNA interference (RNAi) of a target transcript has become a powerful technique to investigate gene function in nematodes and many organisms. Data provided here indicate that the inclusion of 1–2 mM spermidine and 50 mM octopamine and a 24 hr incubation period of nematodes in double-stranded RNA (dsRNA) soaking solutions resulted in a considerable increase in the percentage of nematodes that ingested dsRNA as compared to previous reports. This modified dsRNA soaking method was coupled with quantitative real-time RT-PCR (qRT-PCR) analyses to assess the potential silencing of the Heterodera glycines parasitism gene transcripts Hg-pel-1 and Hg-4E02 that are expressed within the esophageal gland cells of preparasitic H. glycines J2. The Hg-pel-1 transcript was most efficiently silenced with one dsRNA construct (ds267) at the highest dsRNA soaking concentration of 5.0 mg/ml, while the Hg-4E02 transcript was more efficiently silenced at the 2.5 mg/ml dsRNA concentration as compared to 5.0 mg/ml. A dsRNA construct (ds285) complementary to a different sequence within the Hg-pel-1 transcript than construct ds267 induced only minimal silencing of the Hg-pel-1 transcript at 2.5 mg/ml. The results suggest that both dsRNA concentration and sequence relative to the transcript targeted are critical for maximizing potential RNAi effects in parasitic nematodes.

Comments

This article is published as Sukno, Serenella A., Jamie McCuiston, Mui-Yun Wong, Xiaohong Wang, Michael R. Thon, Richard Hussey, Thomas Baum, and Eric Davis. "Quantitative detection of double-stranded RNA-mediated gene silencing of parasitism genes in Heterodera glycines." Journal of nematology 39, no. 2 (2007): 145.

Description
Keywords
Citation
DOI
Source
Copyright
Collections