Evaluation of ploidy variations in Hypericum perforatum L. (St. John’s wort) germplasm from seeds, in vitro germplasm collection, and regenerants from floral cultures

Abstract

Hypericum perforatum L. (St. John’s wort) is an important medicinal herb and a subject of intensive research for its complex and diverse bioactive chemicals. An in vitro-grown germplasm collection of elite H. perforatum lines, established to provide easy access to physiologically uniform plants, was used for ploidy assessment studies. Germplasm lines were maintained by repeated subculture of shoot tips for over 10 yr with little change in their capacity to produce multiple shoots. Shoots of four of these lines were rooted and grown in the greenhouse to obtain plants to provide anther and filament explants. Culture of explants on a regeneration medium supplemented with 1 mg L-1 ?-naphthaleneacetic acid (NAA) and 1 mg L-1 6-benzylaminopurine (BA) induced large numbers of calluses and shoots on all explants. Flow cytometric (FCM) analysis of nuclei samples revealed that the nuclear DNA contents of calluses and shoots developed from anther and filament explants of germplasm lines were not significantly different from those of the donor plants. FCM screening of in vitro-maintained germplasm lines in the collection showed that they had similar nuclear DNA amounts and were all tetraploid (2n = 4x). Analysis of seedlings obtained from the original seed source used to derive the germplasm lines showed that ~11% of them were hexaploid (2n = 6x). Data obtained from FCM screens confirmed the preservation of tetraploidy in in vitro-maintained H. perforatum germplasm and the regenerants obtained from male floral organs. The consistent ploidy of the H. perforatum plants of in vitro origin further supports the usefulness of such technologies to ensure genetic uniformity of medicinal plants over extended periods of culture and may facilitate long-term preservation of their elite clones. © 2015, The Society for In Vitro Biology.