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Rapid recovery- and characterization of transformants following Agrobacterium-mediated T-DNA transfer to sorghum

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Abstract

Although Agrobacterium-mediated transformation of sorghum has been reported, the process is rather lengthy and remains difficult, requiring some very stringent conditions to obtain transformants. We have investigated and describe the parameters related to cocultivation, culture, and regeneration that have allowed us to obtain transgenic sorghum plants in as little as 2.5 months. We observed a 2.9-fold increase in transformation efficiency when L-cysteine was included in the medium during the cocultivation step. Furthermore, the use of modified AB minimal medium, with lower phosphate levels and acidic pH, during the induction of Agrobacterium resulted in a 2.8-fold improvement in transformation efficiencies. Incorporation of an additional binary vector, harboring extra copies of virG and virC genes, in the Agrobacterium did not confer any improvements in the transformation of sorghum. Characterization of transgene activity provided some interesting results suggesting that CaMV 35S promoter activity in T0 generation is very low during the early stages of development of a transgenic sorghum plant, and is not indicative of the expression level during the later stages of development or in the next generation.

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Acknowledgments

We thank Dr. K. Veluthambi for providing us with pSSJ3 plasmid, Dr. G. Sunilkumar for mobilizing it into the A. tumefaciens strain used in this study, and Dr. Venkatachalam Lakshmanan for his help with statistical analysis. We also thank Drs. William L. Rooney and Robert Klein for providing sorghum seeds used in this investigation.

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Authors and Affiliations

  1. Institute for Plant Genomics & Biotechnology, Texas A&M University, College Station, TX, 77843-2123, USA

    Vinod Kumar, LeAnne M. Campbell & Keerti S. Rathore

  2. Department of Soil & Crop Sciences, Texas A&M University, College Station, TX, 77843-2474, USA

    Keerti S. Rathore

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  1. Vinod Kumar
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  2. LeAnne M. Campbell
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  3. Keerti S. Rathore
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Correspondence to Keerti S. Rathore.

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Kumar, V., Campbell, L.M. & Rathore, K.S. Rapid recovery- and characterization of transformants following Agrobacterium-mediated T-DNA transfer to sorghum. Plant Cell Tiss Organ Cult 104, 137–146 (2011). https://doi.org/10.1007/s11240-010-9809-2

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  • DOI: https://doi.org/10.1007/s11240-010-9809-2

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