ARS | Publication request: Micropropagation and Tissue Culture of Hydrangea macrophylla for Mutagenesis, Ploidy Manipulation, and Possible Transformation

Submitted to: American Society of Horticulture Science Meeting
Publication Type: Abstract Only
Publication Acceptance Date: February 2, 2008
Publication Date: February 2, 2008
Citation: Greer, S.P., Reed, S.M., Adkins, J., Rinehart, T.A. 2008. Micropropagation and Tissue Culture of Hydrangea macrophylla for Mutagenesis, Ploidy Manipulation, and Possible Transformation. American Society of Horticulture Science Meeting.

Technical Abstract: We have established parameters for efficient in vitro germination of open pollinated seeds from 17 Hydrangea macrophylla and 2 Hydrangea paniculata cultivars. All tested hydrangea seed had a confirmed light requirement for germination to occur, and contrary to some prior reports, we discovered that germination increased substantially subsequent to cold, wet stratification for one month or greater. In addition, germination was stimulated individually and additively by imbibation with KNO3 and gibberellic acid, as well incubation under low level red light or low level full spectrum light (50-60 footcandles). In contrast, H. macrophylla germination was significantly inhibited by full spectrum light intensities higher than 100 footcandles, KNO3 concentrations higher than 1000ppm, all of our tested concentrations of thiourea, and 6-benzylamino purine (BAP). Using improved germination methods, we have mutagenized seeds from all cultivars using 0.5-5% EMS, assaying seed viability and dormancy of each cultivar with TTC staining before and after treatments. Separate experiments are in progress to develop effectual protocol for the sterile culture, callus and new shoot formation of H. macrophylla explants. Currently, our improved methods establishing sterile cultures from unopened buds have produced success rates approaching 50%; subsequent protocol used to produce regenerative callus and multiple shoots from these sterile cultures has a success rate approaching 10% after 1+ month. We are continuing to evaluate and develop these and additional methods with the goals of transforming and increasing ploidy levels of H. macrophylla.