Regulation of the Bud Dormancy Development and Release in Micropropagated Rhubarb 'Malinowy'.

Culinary rhubarb is a vegetable crop, valued for its stalks, very rich in different natural bioactive ingredients. In commercial rhubarb stalk production, the bud dormancy development and release are crucial processes that determine the yields and quality of stalks. To date, reports on rhubarb bud dormancy regulation, however, are lacking. It is known that dormancy status depends on cultivars. The study aimed to determine the dormancy regulation in a valuable selection of rhubarb 'Malinowy'. Changes in carbohydrate, total phenolic, endogenous hormone levels, and gene expression levels during dormancy development and release were studied in micropropagated rhubarb plantlets. Dormancy developed at high temperature (25.5 °C), and long day. Leaf senescence and dying were consistent with a significant increase in starch, total phenolics, ABA, IAA and SA levels. Five weeks of cooling at 4 °C were sufficient to break dormancy, but rhizomes stored for a longer duration showed faster and more uniformity leaf growing, and higher stalk length. No growth response was observed for non-cooled rhizomes. The low temperature activated carbohydrate and hormone metabolism and signalling in the buds. The increased expression of AMY3, BMY3, SUS3, BGLU17, GAMYB genes were consistent with a decrease in starch and increase in soluble sugars levels during dormancy release. Moreover, some genes (ZEP, ABF2, GASA4, GA2OX8) related to ABA and GA metabolism and signal transduction were activated. The relationship between auxin (IAA, IBA, 5-Cl-IAA), and phenolic, including SA levels and dormancy status was also observed.

Growth Cessation and Dormancy Induction in Micropropagated Plantlets of Rheum rhaponticum 'Raspberry' Influenced by Photoperiod and Temperature.

Dormancy development in micropropagated plantlets at the acclimatization stage and early growth ex vitro is undesirable as it lowers their survival rate and restricts the efficient year-round production of planting material. Thus far, little is known about the factors and mechanisms involved in the dormancy development of micropropagated herbaceous perennials, including rhubarb. This study determined physiological and molecular changes in the Rheum rhaponticum (culinary rhubarb) 'Raspberry' planting material in response to photoperiod and temperature. We found that the rhubarb plantlets that were grown under a 16-h photoperiod (LD) and a temperature within the normal growth range (17-23 °C) showed active growth of leaves and rhizomes and did not develop dormancy. Rapid growth cessation and dormancy development were observed in response to a 10-h photoperiod (SD) or elevated temperature under LD. These morphological changes were accompanied by enhanced abscisic acid (ABA) and starch levels and also the upregulation of various genes involved in carbohydrate synthesis and transport (SUS3, AMY3, BMY3, BGLU17) and ABA synthesis and signaling (ZEP and ABF2). We also found enhanced expression levels of heat shock transcription factors (HSFA2 and HSFA6B), heat shock proteins (HSP22, HSP70.1, HSP90.2 and HSP101) and antioxidant enzymes (PRX12, APX2 and GPX). This may suggest that dormancy induction in micropropagated rhubarb plantlets is a stress response to light deficiency and high temperatures and is endogenously coordinated by the ABA, carbohydrate and ROS pathways.

In Vitro Propagation Method for Production of Phenolic-Rich Planting Material of Culinary Rhubarb 'Malinowy'.

Culinary rhubarb is a popular vegetable crop, valued for its long, thickened stalks, very rich in different natural bioactive ingredients. Tissue cultures are a useful tool for vegetative propagation of virus-free rhubarb plants and rapid multiplication of valuable selected genotypes. The aim of this study was to develop an effective method for in vitro propagation of selected genotypes of Polish rhubarb 'Malinowy' characterized by high yield and straight, thick and intensive red stalks. Identification and quantification of anthocyanins and soluble sugars by the HPLC method in shoot cultures and ex vitro established plantlets were also performed. Shoot cultures were established from axillary buds isolated from dormant, eight-year-old rhizomes. Effective shoot multiplication of rhubarb 'Malinowy' was obtained in the presence of 6.6 µM benzylaminopurine or 12.4 µM meta-topolin. Both cytokinins stimulated shoot formation in a manner that depended on sucrose concentration. Increasing the sucrose concentration from 59 to 175 mM decreased the production of shoots and outgrowth of leaves by 3-fold but enhanced shoot length, single shoot mass and callus formation at the base of shoots. This coincided with increased accumulation of soluble sugars (fructose, glucose) and anthocyanins-cyanidin-3-O-rutinoside (max. 208.2 mg·100 g-1 DM) and cyanidin-3-O-glucoside (max. 47.7 mg·100 g-1 DM). The highest rooting frequency (94.9%) and further successful ex vitro establishment (100%) were observed for shoots that were earlier rooted in vitro in the presence of 4.9 µM indole-3-butyric acid. Our results indicated that anthocyanin contents in leaf petioles were influenced by developmental stage. Under in vitro conditions, it is possible to elicit those pigments by sucrose at high concentration and meta-topolin.