In Vitro Propagation of an Endangered Helianthus verticillatus by Axillary Bud Proliferation

Abstract

Helianthus verticillatus (Asteraceae), whorled sunflower, is a perennial species restricted to a few locations in the Southeastern United States. Habitat loss has caused H. verticillatus to become rare, and since 2014, it has been federally listed as an endangered species. As a part of the recovery plan for the restoration and protection of H. verticillatus, an efficient micropropagation protocol based on axillary shoot proliferation was developed. Various concentrations of 6-benzylaminopurine (BAP; 0 to 4.44 µM) were examined for their morphogenetic potential in the regeneration of six genotypes of H. verticillatus from the nodal explants derived from greenhouse-grown plants. Both the BAP concentration and genotype had significant effects on the regeneration capacity of H. verticillatus. Although the induced buds were observed on ½-strength Murashige and Skoog medium without plant growth regulators, a higher rate of induction and bud development were achieved on media with either 0.88 or 2.22 µM BAP, regardless of the genotype. Successful rooting of the induced shoots was achieved within four weeks after the transfer from the induction medium to the fresh ½-strength MS medium, but the rooting efficiency was dependent on the plant’s genetic background. Regenerated plantlets, with well-developed shoots and roots, were acclimatized successfully to greenhouse conditions with a 97% survival rate. Simple sequence repeats (SSRs) markers were employed to assess the genetic uniformity of the micropropagated plants of H. verticillatus. No extraneous bands were detected between regenerants and their respective donor plants, confirming the genetic fidelity and stability of regenerated plants. To our knowledge, the protocol developed in this study is the first such report for this endangered species.

Genetic diversity and volatile oil components variation in Achillea fragrantissima wild accessions and their regenerated genotypes

“…Some authors reported true-to-mother plant ISSR fingerprinting in the regenerated plants. Examples include Rauwolfia tetraphylla , [ 8 ], Hedychium coronarium [ 7 ], Helianthus verticillatus [ 9 ], and the blackberry [ 10 ]. However, genetic variations of plants in tissue cultures are well reported.…”

“…The genome profiling of plants with pharmaceutical value, using different molecular fingerprinting methods, was reviewed by Gantait et al [ 6 ]. Several of these methods have been applied to address the genetic fidelity of in vitro propagated plants, examples include Hedychium coronarium by inter simple sequence repeats (ISSR) [ 7 ], the endangered medicinal plant Rauwolfia tetraphylla , by start codon targeted polymorphism (SCoT), ISSR, and random amplified polymorphic DNA (RAPD) [ 8 ], Helianthus verticillatus in vitro raised plants using SSRs [ 9 ] and the blackberry ( Rubus fruticosus L.) using RAPD and sequence-related amplified polymorphism (SRAP) [ 10 ].…”

Propagation Methods for the Conservation and Preservation of the Endangered Whorled Sunflower (Helianthus verticillatus)

“…Clonal reproduction methods include various forms of in vitro culture or tissue culture, of which axillary bud proliferation or multiplication is the most common [ 9 ]. Tissue culture or in vitro methods have been applied successfully to the production of H. annuus (common sunflower) plants e.g., [ 10 ] as well as to H. verticillatus [ 11 ]. Typically, axillary bud proliferation [ 9 ] involves excising quiescent axillary buds and a small segment of the stem or a node (together, termed an explant) from an elite plant to be cloned.…”

Callus-Mediated High-Frequency Plant Regeneration, Phytochemical Profiling, Antioxidant Activity and Genetic Stability in Ruta chalepensis L.

“…However, synergistic interactions between BA (5.0 µM) and NAA (1.0 µM) resulted in a greater response in the induction and multiplication of shoot buds from the callus mass. The optimal amount of cytokinins and auxins in the medium is critical for the induction and multiplication of shoot buds since it varies depending on the kind of explant, the cultivar, and the type of plant [36]. In their study, Wang, et al [37] found that excessive cytokinins and auxins adversely influence plant regeneration.…”