In this study, a tissue culture method was developed for the propagation and conservation of Cattleya gaskelliana (N.E.Br.) B.S. Williams. Initially, protocorms with leaf primordium, obtained via asymbiotic seed germination, were used as explants. Half-strength Murashige and Skoog (MS) medium supplemented with 1.5 mg L⁻¹ 6-benzyl aminopurine (BA) and 2.0 mg L⁻¹, indole butyric acid (IBA) was found to be optimal for the proliferation of protocorm-like bodies (PLB). Most explants (81%) proliferated, each producing 11.1 ± 1.87 units per explant. The optimum number of PLB differentiated to plantlets on a cultured medium with 0.5 mg L⁻¹ Thidiazuron (TDZ) + 100 mL L⁻¹ coconut water. Firmly rooted plantlets with vigorous growth were regenerated on ½ MS medium + 1.0 mg L⁻¹ Indole- 3-acetic acid (IAA) and 50 g L⁻¹ banana puree. Plants regenerated via in vitro processes were wrapped with sphagnum moss and acclimatized in a substrate containing equal proportions of bark, perlite, and peat with a 100% ex-vitro survival rate after acclimation for 70 days. The protocol developed in this study can be used to obtain several thousand plants within one year, and thus, is an effective method.

Trichopilia suavis is one of the important representatives of the genus Trichopilia, which can be used as a potted plant for the interior design. also, its beautiful inflorescences are added unusual element to bouquet compositions and actively used in the production of perfumes. This study was carried out in order to develop and improve micropropagation method for mass clonal production of T. suavis through protocorm culture. Half-strength Murashige and Skoog culture medium (½) has been modified by adding different organic components and growth regulators. The protocorms obtained from seeds germinated by asymbiotic method were used as explants. The results showed that the optimal treatment for formation of new protocorms (7.50 ± 0.7 units per protocorm) was obtained by culture on ½ MS medium with 0.5 mg L^-1 of 6-Benzylaminopurine (6-BAP) and 100 mL L^-1 of coconut water. At the stage of rooting, the highest number and length of roots (7.0 ± 0.45 units per plantlet, 2.64 ± 0.18 cm) were obtained through subsequent cultivation of plantlets on ½ MS medium with 0.5 mg L^-1 indolyl-3-butyric acid (IBA), 1.0 g L^-1 charcoal with the addition of 50 g L^-1 of banana puree. The obtained seedlings were successfully adapted in a substrate consisting of bark, perlite and peat in a ratio of 1:1:1.

Cymbidium erythrostylum Rolfe is one of the most beautiful species of the genus cymbidium which is used as a cut flower or indoor plant. However, it was registered as a rare species even in its original habitat. This study was carried out to develop a protocol for in vitro propagation of C. erythrostilum. We used protosomes obtained by the asymbiotic germination of seed on 1/2 of Murashige and Skoog nutrient medium (MS) supplemented with 1 mg/L 2-Isopentenyl adenine (2iP) as objects of study. During the multiplication stage, the number of formed protosomes on a culture medium containing 0.8 mg/L 6-Benzylaminopurine (6-BAP) was higher (3.8 ± 0.2 protosomes). During the subsequent cultivation on ½ MS medium supplemented with 2 mg/L 6-BAP and 0.5 mg/L a-naphthaleneacetic acid (NAA), the highest numbers of shoots (4.00 ± 0.19 shoots/plantlet) and leaves (4.50 ± 0.14 leaves/plantlet) were obtained. At the rooting stage under in vitro conditions, the most effective was the use of ½ MS nutrient medium with the addition of 0.5 mg/L of indolyl-3-butyric acid IBA, 1 g/l charcoal, and 50 g/l banana puree. The obtained plants were successfully adapted to a substrate consisting of bark, perlite, and peat in a ratio of 1:1:1.

Dactylorhiza fuchsii (Druce) Soó is a rare species listed in the Red Books of the Moscow Region and 25 regions of the Russian Federation. This study focuses on optimizing clonal micropropagation and conservation methods for D. fuchsii and identifying anatomical features of underground organs under in vitro and ex vitro conditions. The results showed that cultivating protocorms on 1/2 MS medium with 1.5 mg/L 6‑BAP and 1.5 mg/L IBA under light conditions is optimal for forming new protocorms. Conversely, 1/2 MS medium with 1.5 mg/L 6-BAP and 2.0 mg/L IBA under dark conditions promoted protocorm development and seedling formation. For seedling development, 1/2 MS medium with 1.0 mg/L 6-BAP and 100 mL/L coconut water was most effective. The highest numbers of protocorm-like bodies were obtained with 1/2 MS medium containing 2.0 mg/L KIN and 20 g/L potato homogenate. Seedlings were successfully adapted after three months of cold treatment at 5°C in the dark, followed by growth in a substrate of bark, perlite, sand, and peat moss. Anatomical analysis showed that increased root diameter results from an increase in the number of parenchyma layers and cell size, as well as an increase in the number and development of vascular bundles in the stele. Tuber anatomy was also studied for the first time under in vitro and ex vitro conditions. This study provides valuable insights for the large-scale propagation of this endangered species and their reintroduction into the environment to preserve biodiversity.

Phragmipedium kovachii is a renowned orchid for its striking flowers and ecological significance. This study aimed to develop an efficient protocol for its micropropagation and conservation. Shoots measuring 0.7-1.0 cm were used as explants and cultured on half-strength Murashige and Skoog (MS) medium with various combinations of 6-benzylaminopurine (6-BAP) and indole-3-acetic acid (IAA), with or without 100 mL L^-¹ coconut water. The highest protocorm-like bodies (PLBs) formation rate (96.7%) and the maximum PLBs per explant (12.58 ± 0.33) were obtained on medium containing 2.0 mg L^-¹ 6-BAP, 0.6 mg L^-¹ IAA, and coconut water. During the regeneration phase, PLBs regenerated on media supplemented with either 0.5 mg L^-¹ 6-BAP and coconut water, as well as those on media containing 0.5 mg L^-¹ meta-Topolin (mT) and banana homogenate, produced optimal plantlets for subsequent rooting stages. Rooting was enhanced using 1.0 mg L^-¹ IAA with coconut water or 0.5 mg L^-¹ indole-3-butyric acid (IBA) with banana homogenate. Plantlets grown on coconut water-supplemented media developed longer roots with a pentarch stele, well-differentiated sclerenchyma tissue, and numerous metaxylem vessels, exhibiting the highest survival rate. In contrast, those cultured on banana homogenate produced thicker roots with a polyarch stele. Acclimatization was successfully achieved using substrates consisting of equal proportions of bark, perlite, and peat, as well as a mixture of soil, sand and perlite (2:1:1), ensuring high ex vitro survival rates (87 and 80%, respectively). This study provides an effective protocol for the ex situ conservation and large-scale production of P. kovachii plants.

The present article is devoted to optimizing rooting of Cattleya gaskelliana regenerants in vitro, and to identifying anatomical features of root under various culture conditions in vitro. The results showed that semi-solid 1/2 MS nutrient medium containing 50.0 g/L banana puree and 1.0 mg/L IAA is more effective in the growth and rooting of C. gaskelliana regenerants. Also, it has been determined the histological differentiation of C. gaskelliana roots formed on nutrient media with different concentrations of agar. Plants cultured on semi-solid nutrient medium had roots with more developed diameter and a central cylinder. To obtain maximum survival rate (100%), it is recommended to use sphagnum moss and a substrate consisting of bark, perlite, and peat in equal ratio.