Ingestion of Double-Stranded RNA by Preparasitic Juvenile Cyst Nematodes Leads to RNA Interference
Tóm tắt
RNA interference is of value in determining gene function in many organisms. Plant parasitic nematodes are refractory to microinjection as a means of introducing RNA and do not show any oral uptake until they are within plants. We have used octopamine to stimulate uptake by prepara-sitic second stage juveniles of two cyst nematodes, Heterodera glycines and Globodera pallida. This new technique was used to facilitate uptake of double stranded RNA (dsRNA) together with fluoroscein isothiocyanate as a visual marker. Targeting cysteine proteinases did not reduce the number of parasites but caused a shift from the normal female/male ratio of 3:1 to 1:1 by 14 days postinfection (dpi). Exposure of H. glycines to dsRNA corresponding to a newly characterized protein with homology to C-type lectins did not affect sexual fate, but 41% fewer parasites were recovered from the plants. As expected, treatment with dsRNA corresponding to the major sperm protein (MSP) had no effect on either parasite development or sexual fate over 14 days. Northern analysis showed lower transcript abundance for the two targeted mRNAs that occur in J2, plus a later inhibition for MSP transcripts when males developed sperm at 15 dpi. These findings establish a procedure for RNAi of plant parasitic nematodes.
Từ khóa
Tài liệu tham khảo
Atkinson H. J., 1996, J. Nematol., 28, 209
Atkinson, H. J., Winter, M. D., and McPherson, M. J. September 2000. Control of crop pests and animal parasites through direct neuronal uptake. U.S. patent 46070GB
Kamath, R. S., Martinez-Campos, M., Zipperlen, P., Raser, A. G., and Ahringer, J. 2000. Effectiveness of specific RNA-mediated interference through ingested double-stranded RNA in Caenorhabditis elegans. Genome Biol. 2:research0002.1
Novitski C. E., 1993, J. Nematol., 25, 548
Sambrook, J., Fritsch, E. F., and Maniatis, T. 1989. Molecular Cloning: A Laboratory Manual. Cold Spring Harbor Laboratory Press. Cold Spring Harbor, NY, U.S.A.
Sasser, J. N., and Freckman, D. W. 1987. A world perspective on nematology: The role of the society. Pages 7-14 in: Vistas on Nematology. J. A. Veech and D. W. Dickerson, eds. Society of Nematologists. Hyattsville, MD, U.S.A.
Urwin P. E., 1997, Parasitology, 114, 605
Willats W. G. T., 1995, J. Nematol., 27, 135
Wyss U., 1992, Fundam. Appl. Nematol., 15, 75