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Genetic transformation of cultivated sesame (Sesamum indicum L. cv Rama) through particle bombardment using 5-day-old apical, meristematic tissues of germinating seedlings

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Abstract

An in vitro plant generation and genetic transformation protocol was established in sesame (Sesamum indicum L. cv Rama) through biolistic particle gun bombardment. 5-day-old apical, meristematic tissues of in vitro-germinating seedlings were used as explants. 10–15 Multiple shoots were generated from each explant using Murashige and Skoog basal medium containing 18.0 µM benzylamino purine and 5.37 µM naphthalene acetic acid. Four independent sets of transformation were carried out and each set consisted of three independent experiments each comprising three replications with 30 explants per replication. A synthetically designed bialaphos resistance gene (bar) was used for transformation. The positive transformants containing the bar gene were selected in growth medium containing 2.5 mg/L bialaphos. Green shoots recovered from bombarded explants were subjected to root development on Murashige and Skoog basal medium containing 5.37 µM naphthalene acetic acid. The rooted shoots were established in soil and grown to maturity in greenhouse. Polymerase chain reaction (PCR), Southern and reverse-transcription PCR, real-time quantitative PCR, western blot and enzymatic assay of four putative transformants from independent sets provided evidence for full-length gene integration as well as high level expression of the transgene. Analysis of the T1 plants revealed a stable inheritance of the transgene through the progenies. This is the first report of biolistic mediated stable transformation of sesame and should pave the way for future genetic engineering strategies to be employed for improvement of this very important oil-seed crop.

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Acknowledgments

The authors extend their sincere appreciation for Shila Bhattacharjee for her extensive assistance in the tissue culture. The authors are also thankful to Sudarshan Maity, Uttam Dogra and Subhash Ghosh for maintenance and rearing of transgenic sesame plants under containment conditions. Grant support to the laboratory from Indian Council of Agricultural Research (ICAR) is thankfully acknowledged. Finally, the authors express their sincere gratitude to the distinguished editor and the anonymous reviewers, whose incisive comments were of immense help for improvement of the quality of the manuscript.

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Correspondence to Soumitra Kumar Sen.

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Bhattacharyya, J., Chakraborty, A., Mitra, J. et al. Genetic transformation of cultivated sesame (Sesamum indicum L. cv Rama) through particle bombardment using 5-day-old apical, meristematic tissues of germinating seedlings. Plant Cell Tiss Organ Cult 123, 455–466 (2015). https://doi.org/10.1007/s11240-015-0848-6

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