Agrobacterium-mediated transformation in Darwin’s Gloxinia, Sinningia speciosa
Date Issued
2012
Date
2012
Author(s)
Chan, Zuo-Hao
Abstract
Sinningia speciosa is an ornamental plant species that is widely cultivated. Many cultivars, however, generated a range of floral mutations such as Peloria (actinomorphy). It thus allows us to examine candidate genes that control floral symmetry. These functions should be able to be clarified if a suitable transformation system is established. Although there was a transformation protocol published on S. speciosa, yet it mainly focused on genetic analysis. Due to this reason, a more detailed transformation should be clarified. We therefore chose Agrobacterium, strand LBA4404, with medium Hygromycin as a selective agent, to infect leaf explants after they were sectioned into the size of 0.5 cm2. We also found out that S. speciosa has the capability to regenerate vigorously through vegetative cuttings after being placed on MS medium containing 2 ppm of BA and 0.2 ppm of NAA. So the major optimizing factor is on Agrobacterium infections. New plantlets induced from shoots were beta-glucuronidase(GUS)-stained and confirmed with gDNA PCR and RT-PCR in order to predict the rate of success. Out of all 120 explants, 14 regenerants were collected and five of which was successfully transformed as confirmed by PCR using GUS primers, while all of which showed GUS expression as confirmed by RT-PCR, resulting in a transformation rate of 4.16%. This project shall serve as an important mile stone in building a reliable transformation protocol in Gesneriaceae, allowing functional validations of several candidate floral developmental genes in evo-devo, thus assists any possible future project which could be brought out by our laboratory.
Subjects
Agrobacterium-mediated transformation
Sinningia speciosa
Gesneriaceae
GUS staining
Tissue culture
Type
thesis
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