In Vitro Segregation of Tetraploid and Octoploid Plantlets from Colchicine-induced Ploidy Chimeras in Echinacea purpurea L.

Echinacea purpurea L. is one of the important ornamental and medicinal plant species. Ploidy manipulation is a valuable tool for improving plant quality or production in E. purpurea as well as in many other plants. To study the segregation of pure ploidy plantlets from colchicine-induced ploidy chimeras in E. purpurea, we used a chimera plantlet that consisted of 1.93% diploid, 35.04% tetraploid, and 63.03% octoploid cells as the source material for experiments. The results showed that three factors significantly influenced the segregation, i.e., the component ratios of different ploidy cells in the chimera, the number of sequential passages, and the methods of segregation culture of the chimera plantlets. Other factors, such as explant types (i.e., leaf, petiole, or root) and 6-benzyladenine (BA) concentrations (i.e., 0.2, 0.4, 0.8, and 1.2 mg·L−1) occasionally influenced the segregation. Pure chromosome-doubled polyploids are not easily obtained in various plant species, so segregation culture of ploidy chimeras may potentially be more effective. The morphological characteristic and content of cichoric acid were compared among diploid, tetraploid, and octoploid plants. Results indicated that tetraploid and octoploid plants had more stunted growth, larger stomata, lower stomata frequency, more chloroplast number in guard cells, and higher cichoric acid content than original diploid lines.

Laboratory Techniques for Determining Ploidy Levels

The determination of ploidy levels is essential for breeding asexually propagated and polyploid crops that may have ploidy chimeras in their tissues or have several polyploid series. This presentation will discuss a laboratory for teaching students how to determine ploidy levels in different plant tissues using different techniques. The different methods for ploidy determination include root tip squashes, pollen mother cell squashes, pollen grain size and germinal pore counts, stomata size and density determination, and gross morphology. After completing these laboratory experiments, students will be able 1) to initiate a study of ploidy determination, 2) to determine the correct course of action in determining ploidy level, and 3) to recognize the various steps required to determine ploidy level in plants.

Colchicine-induced Amphidiploids of Rose Interspecific Hybrids

An objective of our rose breeding research is to transfer resistance to blackspot and other diseases from wild diploid species to modern rose cultivars. Interspecific hybrids among blackspot-resistant diploid species were chosen for chromosome doubling to produce fertile amphidiploids that could be hybridized to the tetraploid commercial germplasm. Five such F1 interspecific hybrids were treated with colchicine. The study included two different application procedures (shake in colchicine solution or colchicine in media), four colchicine concentrations (0.05%, 0.1%, 0.15%, and 0.20%), and five treatment periods (1, 3, 5, 8, and 10 days). After colchicine treatment, all the materials were cultured in vitro. One thousand-thirty-seven surviving explants were selected for typical “gigas” characteristics of doubled diploids. Chromosome counts on shoot tips of these selected genotypes confirmed 15 amphidiploids. The best colchicine treatment varied among the interspecific hybrids. Higher colchicine concentrations or duration reduced growth rating, rooting, and percent survival. The recognition of amphiploids and ploidy chimeras from young seedlings will also be discussed.