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United States Department of Agriculture

Agricultural Research Service

Title: High Efficiency Induction of Soybean Hairy Roots and Propagation of the Soybean Cyst Nematode

Authors
item Cho, Hyeon-Je - U OF ILL, URBANA
item Farrand, Stephen - U OF ILL, URBANA
item Noel, Gregory
item Widholm, Jack - U OF ILL, URBANA

Submitted to: Planta
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: August 8, 1999
Publication Date: N/A

Interpretive Summary: Soybean cyst nematode (SCN), Heterodera glycines, is a serious pest of soybean throughout soybean production areas in the United States, causing an estimated annual production loss of 80 million bushels or $400 million. A major component of SCN control is the use of nematode resistant soybean. Planting of resistant varieties can be used to minimize crop loss to the soybean producer. However, unless managed carefully, resistance to SCN may not be durable. That is, new races of SCN can be selected that can overcome resistance that is available at present. Domesticated soybean has a small number of genes that confer resistance to SCN and nematode populations exist that can overcome all known sources of resistance in domesticated soybeans. Biotechnology shows promise in providing researchers with the ability to incorporate exotic resistance genes into domesticated soybean. The research reported herein concerns the efficient production of genetically altered soybean roots that can be used to study novel genes and their expression in the genetically engineered soybean roots. The ability to express novel genes such as the green fluorescent pigment gene and subsequent culture of SCN on the genetically engineered soybean roots, shows great promise in being able to transfer novel genes from various organisms into domesticated soybean and to test the resistance of these genes to SCN rapidly under laboratory conditions. All the resistance genes in soybean have been utilized and can be overcome by SCN. Soybean producers will benefit from new types of resistance from other crops and organisms as these genes for which SCN has not evolved resistance are moved into soybean. Thus, resistance in genetically modified soybean will last longer and increase soybean production efficiency and profit.

Technical Abstract: Cotyledon explants of 10 soybean cultivars were inoculated with Agrobacterium rhizogenes strain K599 with and without binary vectors pBI121 or pBINm-gfp5-ER possessing both nptII and gus or nptII and gfp genes, respectively. Hairy roots were produced from the wounded surface of 54-95% of the cotyledon explants on MXB selective medium containing 200 ug ml**-1 kanamycin and 500 ug ml**-1 carbenicillin. Putative individual transformed hairy roots were identified by cucumopine analysis and were screened for transgene incorporation using PCR. All of the roots tested were found to be co-transformed with T-DNA from the Ri-plasmid and the transgene from the binary vectors. Southern blot analysis confirmed the presence of the 35S-gfp5 gene in the plant genomes. Transgene expression was also confirmed by histochemical GUS assay and Western blot analysis for the GFP. Attempts to induce shoot formation from the hairy roots failed. Infection of hairy roots of the soybean cyst nematode Heterodera glycines, susceptible cultivar, Williams 82, with eggs of H. glycines race 1, resulted in the development of mature cysts about 4-5 weeks after inoculation. Thus the soybean cyst nematode could complete its entire life cycle in transformed soybean hairy root cultures expressing GFP. This system should be ideal for testing genes that might impart soybean cyst nematode resistance.

Last Modified: 12/17/2014
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