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A simple and efficient method for obtaining transgenic soybean callus tissues

Liu, Dong, Liu, Sheng, Chang, Dasong, Wang, Lu, Wang, Dan, Wang, Ning Ning
Acta physiologiae plantarum 2013 v.35 no.7 pp. 2113-2125
2,4-D, Agrobacterium, Glycine max, antioxidants, benzyladenine, callus, explants, genes, genotype, hypocotyls, phenotype, screening, soybeans
In the present study, a simple and efficient method for obtaining transgenic callus tissues of soybean [Glycine max (L.) Merr.] was developed based on Agrobacterium-mediated transformation. Hypocotyl segments of soybean were used as the starting material. Several factors such as soybean genotype, Agrobacterium concentration, inoculation time, co-cultivation period and addition of antioxidants in co-cultivation medium affecting the transformation efficiency were examined. The explants were cultured on callus induction medium containing 0.5 mg L⁻¹6-benzylaminopurine and 2.0 mg L⁻¹, 2,4-Dichlorophenoxyacetic acid for callus induction. Callus tissues were induced at both the acropetal and basipetal ends. CaMV35S::GUS and CaMV35S::GFP transgenic callus tissues were obtained using the optimized protocol. The average transformation efficiency reached up to 87.7 % based on GUS detection. From inoculation with Agrobacterium to obtaining transgenic soybean callus will take about 3 weeks. In order to validate this method for gene function investigation, GVG::GmSARK transgenic soybean callus tissues were obtained and their senescence-associated phenotypes were assessed. To our knowledge, this is the first report using hypocotyl segments as starting materials to obtain transgenic callus, and this system provides a method for high-throughput screening of functional genes of interest in transformed soybean callus.