Modification of plant regeneration medium decreases the time for recovery of Solanum lycopersicum cultivar M82 stable transgenic lines
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Tomato (Solanum lycopersicum) has rapidly become a valuable model species for a variety of studies including functional genomics. A high-throughput method to obtain transgenic lines sooner than standard methods would greatly advance gene function studies. The goal of this study was to optimize our current transformation method by investigating medium components that would result in a decreased time for recovery of transgenics. For this study, 6-day-old cotyledon explants from Solanum lycopersicum cultivar M82 in vitro-grown seedlings were infected with the Agrobacterium tumefaciens strain LBA4404 containing the binary vector pBI121. This vector contains the β-glucuronidase reporter gene and the neomycin phosphotransferase II selectable marker gene that confers resistance to kanamycin. Modification of our standard plant regeneration medium with indole-3-acetic acid (IAA) at concentrations of either 0.05 or 0.1 mg/l decreased the recovery time for transgenic lines by 6 weeks as compared to our standard medium that contains zeatin as the only plant growth regulator. We observed 50 and 54 % transformation efficiency on plant regeneration medium containing 0.05 and 0.1 mg/l IAA, respectively. Moreover, addition of 1 mg/l IAA to the root induction medium resulted in earlier root development than medium that did not contain IAA. Addition of IAA to the plant regeneration and rooting media did not have any negative effects on plant development. Recovery of transgenic lines in a shorter time results in higher throughput for the introduction of gene constructs and has the potential to decrease the time and resources needed to complete investigations of gene function.
KeywordsAgrobacterium tumefaciens Indole-3-acetic acid Solanaceae Solanum pimpinellifolium Tomato
We thank Cynthia Du for assistance with part of the experimental process. We thank Cynthia Du, Patricia Keen, and Michelle Tjahjadi for their critical review of the manuscript. Support for this work was through a grant from the National Science Foundation Plant Genome Research Program (IOS-1237880).
SG and JVE designed the experiments, SG performed the experiments, SG and JVE wrote the manuscript. Both authors read and approved the manuscript.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no competing interests.
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