Abstract
Few reports explain the mechanism of PEG action on stomatal behavior and anatomical structure and analyze the photosynthetic pigments of in vitro date palm plantlets for better tolerance to ex vitro exposure. The main challenge for in vitro micropropagation of date palm techniques remains restricted to high survival rates and vigorous growth after ex vitro transplantation. In vitro hardening is induced by Polyethylene glycol PEG (0.0, 10, 20, 30 g L−1) for 4 weeks. Leaf anatomy, stomatal behavior, water loss %, photosynthetic pigments, and reducing sugars were examined in date palm plantlets (Phoenix dactylifera L.) cv. (Sewi) after 4 weeks from in vitro PEG treatment and after 4 weeks from ex vitro transplanting to the greenhouse. Leaf anatomy and the surface ultrastructure of in vitro untreated leaves showed a thin cuticle layer, wide opened malfunctioning stomata, and abnormal leaf anatomy. Furthermore, addition of PEG resulted in increasing cuticle thickness, epicuticular wax depositions, and plastids density, improving the stomatal ability to close and decreasing the stomatal aperture length while reducing the substomatal chambers and intercellular spaces in the mesophyll. As a result, a significant reduction in water loss % was observed in both in vitro and ex vitro PEG treated leaves as compared to untreated ones, which exhibited rapid wilting when exposed to low humidity for 4 h. PEG application significantly increased Chlorophylls a, b and carotenoids concentrations, especially 10, 20 g L−1 treatments, which were sequentially reflected in increasing the reducing sugar concentration. However, leaves of plantlets treated with PEG at 30 g L−1 became yellow and had necrosis ends with death. In vitro hardening by 20 g L−1 PEG increased the survival rate of plantlets to 90% after ex vitro transfer compared to 63% recorded for the untreated plantlets. Therefore, this application provides normal date palm plantlets developed faster and enhances survival after ex vitro transfer.
Highlights
Date palm (Phoenix dactylifera L.) is one of the most important fruit trees in arid and semiarid regions of the Middle East and North Africa [1]
The lack or poor formation of the cuticle is common in plants grown under in vitro conditions [30], where high relative humidity inside the culture vessels represses cuticle formation [24]. This layer is an important factor for surviving the in vitro plantlets during transplantation to the ex vitro conditions, via helping in control transpiration passive water loss [31]. This may explain the increase in water loss rates in both in vitro and ex vitro untreated leaves compared to the PEG treated plantlets after 4 h of air drying treatment
The results indicated that the percentage of water loss in the in vitro control detached leaves was higher than that recorded in the ex vitro control ones (46.13 and 41.24%, respectively, Table 3)
Summary
Date palm (Phoenix dactylifera L.) is one of the most important fruit trees in arid and semiarid regions of the Middle East and North Africa [1]. Palm trees have been planted by separating the offshoots and independently established in the fields. This traditional method is not sufficient due to the limited number of offshoots produced from the mother tree in its lifespan. In vitro micropropagation techniques became effective methods for rapid and mass multiplication by either somatic embryogenesis [3,4] or organogenesis protocols [5,6]. The success of these protocols on a commercial scale is highly dependent on the ability of regenerated plantlets to transfer from in vitro controlled condittions and well established in the newly hard ex vitro conditions
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