Cyrtopodium paranaense is a tropical terrestrial orchid, which propagates mainly through sexual seed germination. In this study, we document the asexual morphogenesis of the root tip to protocorm-like body (PLB) conversion in Cyrtopodium paranaense . Protocorm-like bodies sporadically developed from root tips of flask-grown seedlings in the absence of any exogenous plant growth regulators (PGRs). The compact PLBs ultimately gave rise to normal plantlets. Histological observations revealed that the root cap became dissociated from the root apex at an early stage followed by dispersed extension of root vascular strands into nascent PLBs. Protocorm-like bodies also developed from the root central stele tissue. In root tip segment cultures, PLBs were not formed without providing PGRs but were efficiently formed from root tips in Murashige and Skoog (MS) medium supplemented with 10.2 μM indole-3-acetic acid (IAA) and 9.0 μM thidiazuron (TDZ). Both IAA and TDZ promoted the formation of PLBs; however, TDZ did not induce PLB formation in the absence of IAA, indicating a synergistic effect of the two PGRs. Protocorm-like bodies were proliferated and subsequently plants regenerated in PGR-free MS medium. Root tip culture may be used as an alternative method for the propagation of Cyrtopodium paranaense .

This first-attempt study used microtome-based methods to generate a thin cell layer culture for the micropropagation of Phal. Hwafeng Redjewel × Phal. New Cinderella. Protocorms were embedded in various agarose concentrations (8–12%, w/v) and dried from 1 to 8 h before sectioning with a microtome. Optimal conditions for slicing sections of 100 to 300 μm were achieved when the protocorms were embedded at 10% (w/v) agarose and dried for 4 h under laminar flow, and the hardness of the agarose block under these conditions reached 641.8 ± 9.5 g·cm⁻². The sectioned protocorms that were cultured on an MS medium supplemented with 1.2 mg·L⁻¹ 6-benzylaminopurine and 0.1 mg·L⁻¹ α-naphthaleneacetic acid were capable of growth and differentiated through the neoformation of protocorm-like bodies (PLBs) and/or callus before subsequent regeneration into plantlets and development into healthy plants in a nursery environment. The results of this study demonstrate that microtome-based tTCL is a reliable and promising approach for mass propagation and possible virus-free propagation objectives for Phalaenopsis.

Background: Adiantum reniforme var. sinense is a pteridophyte with considerable ornamental importance and medicinal value. However, the wild population of A. reniforme var. sinense has gradually become scarce due to sparse distribution and human activities. Green Globular Bodies (GGBs) are specific tissue which use to propagate ferns in vitro. The study uses GGBs induced from sporophytic embryos to preserve and to proliferate this pteridophyte. Methods: The present study was carried out during year 2018-2020 at National Ilan University. The developing sporophyte of Adiantum reniforme var. sinense were used to induce GGBs. The optimal medium for GGBs proliferation and differentiation in terms of basal salts, plant growth regulators, sugar concentrations and gelling agents was examined. The structure of GGBs and the embryos were analyzed by paraffin section as well. Result: The result indicates the 100% GGBs induction rate from sporophytic embryos for the first time could be obtained under BA (0.1 mg L-1) and NAA (0.15 mg L-1). For subsequent proliferation of GGBs required a higher concentration of NAA (10 mg L-1). Cytokinins such BA, KT and TDZ, inhibited GGBs proliferation. In vitro optimal amount of basal salts, plant growth regulators, sugar concentrations and gelling agents for GGB proliferation were obtained as 2 g L-1 Hyponex, 10 mg L-1 NAA, 15 g L-1 sucrose and 4 g L-1 gelrite, respectively. Paraffin sections revealed histological characteristics of the sporophytic embryo and GGB.

The induction of green globular bodies (GGBs) represents an unique phenomenon in the tissue culture of pteridophytes. In this first attempt study GGBs were induced with the embryos of Adiantum reniforme var. sinense. The influences of basal salts, plant growth regulators, sugar concentrations, and gelling agents on the proliferation and differentiation of GGBs were examined. The highest induction rate of GGBs resulted from Murashige & Skoog (MS) medium supplemented with benzyl aminopurine (BA) (0.1 mg L⁻¹) and 1-naphthaleneacetic acid (NAA) (0.15 mg L⁻¹). The subsequent proliferation of GGBs required a higher concentration of NAA (10 mg L⁻¹) to have the highest fresh weight of 0.29 g. However, cytokinins, such as BA, kinetin (KT), and thidiazuron (TDZ), exhibited inhibitory effects on GGB proliferation. Hyponex was superior to MS medium in terms of GGBs proliferation and differentiation, which led to the highest fresh weight (1.72±0.63 g) and shoot number (7.25±1.89) after 60 days of cultivation. Among different sucrose concentration in media, GGBs cultured with 15 g L⁻¹ sucrose had the highest fresh weight (1.01 g). Gelling agents also showed evident effects on the proliferation of GGBs, and the fresh weight of GGBs cultured in gelrite medium was approximately three times greater than that of GGBs cultured in agar medium. Distinct histological characteristics of embryos and GGBs were revealed by paraffin sections. All regenerates from GGBs survived in the greenhouse, with normal appearances, after acclimation in the medium of peat soil and pearlite (1:1).

The green globular body is a unique, organized tissue that emerges from explants in pteridophyte tissue cultures. This article describes the first comprehensive study of in vitro regeneration of Adiantum trapeziforme via green globular bodies. Green globular bodies were induced on the rhizomes of sterile sporophytes of Adiantum trapeziforme . The green globular bodies were used as explants for additional proliferation and regeneration. The study evaluated the concentrations of basal salts, sucrose, tryptone, and plant growth regulators. The results showed that the presence of 6-benzyladenine inhibited the proliferation and regeneration of green globular bodies, whereas while increasing the concentration of α-naphthaleneacetic acid to 5 mg·L ⁻¹ led to the highest proliferation and drastically impeded the development of green globular bodies. Hyponex No. 1 fertilizer, especially at 3 g·L ⁻¹ , with a proliferation rate up to 8.8 times, was a better salt source than Murashige and Skoog medium. In the sucrose concentration test, 30 g·L ⁻¹ sucrose resulted in the highest proliferation rate. Furthermore, medium with a higher tryptone concentration inhibited the proliferation of green globular bodies. Green globular body differentiation medium containing 2 g·L ⁻¹ Hyponex, 30 g·L ⁻¹ sucrose, 2 g·L ⁻¹ tryptone, and pH 5.8 resulted in the highest bud induction rate, averaging 18.6 buds. Paraffin sections were examined to observe the development of green globular bodies induced by the rhizome. In the soil containing perlite, 100% of regenerated shoots successfully formed roots during acclimation.