Gas exchange measurement as a non-destructive viability assay for frozen-thawed, winter-dormant apple buds.

Low temperature studies with winter-dormant buds are severely limited by the lack of a rapid, non-destructive assay for their viability. Investigations involving the winter harvest of ecodormant buds of woody subjects, including cryopreservation, are restricted if viability cannot be assessed until dormancy is broken. If post-treatment grafting indicates low survival of the harvested population then further collection and study has to be delayed until the next winter season. This study trials the use of a portable gas exchange system able to discriminate between live and dead buds rapidly, with the assay confirmed as non-destructive by subsequent micropropagation. Active respiration was recorded for 85% of a winter-dormant Malus domestica buds population that showed 91% viability when grafted (n = 45). Lethally stressed material gave no false positive results. When micropropagated after respiratory measurement, a population viability of 76% was recorded. There was a significant, positive correlation between respiration and fresh weight for buds of mass >10 mg, from a population with a mean fresh weight of 17 mg.

Determining the effect of pretreatments on freeze resistance and survival of cryopreserved temperate fruit tree dormant buds.

Freeze resistance is critical to successful dormant bud (DB) cryopreservation, and is affected by genotype, environmental conditions, dormancy phase and processing techniques. Pretreatment induced freeze resistance may contribute to more successful and efficient protocols for cryopreserving DB. Differential thermal analysis (DTA) was used to quantify the effects of cryopreservation pretreatments on freeze resistance of dormant budwood. Low temperature exotherm (LTE) profiles created by DTA could rapidly identify pretreatments that are contributing to increased freeze resistance in tree fruit species. In this study, DTA was used to help elucidate the effects of varying pretreatments (sucrose, desiccation and their combination) on peach, a model crop in tree fruit physiology that has shown little cryosurvival using the DB method in the past. Post cryopreservation recovery trials using an antimicrobial forced bud development (AFBD) protocol evaluated the ability of selected pretreatments, that improved freeze resistance based on DTA, to improve recovery of dormant budwood of various deciduous tree fruit and nut species. Precryogenic exposure to sucrose solution (5.0 M, 96 h), desiccation to 30% moisture content (MC) and their combination tested for their efficacy on improving postcryogenic viability in peach, apricot, sweet cherry, little walnut, black walnut, English walnut, apple, and pear. Among the different pretreatments tested, desiccation to 30% MC had the greatest impact on increasing freeze resistance and cryosurvival across most fruit species tested and little walnut. Gradual reduction of MC (from 40 to 25%) levels increased freeze resistance in peach (R2=0.95) and increased some recovery outcomes (leaf, shoot and bud swell), however, this was not correlated with equal cryorecovery outcomes as severe bud cracking was observed. Overall, our approach linking freeze resistance and preconditioning treatments could help establish efficient species-specific cryopreservation protocols for a number of important temperate woody crops which could be recovered as complete plants by coupling AFBD and plant tissue culture.

A device for the controlled cooling and freezing of excised plant specimens during magnetic resonance imaging.

BACKGROUND: Investigating plant mechanisms to tolerate freezing temperatures is critical to developing crops with superior cold hardiness. However, the lack of imaging methods that allow the visualization of freezing events in complex plant tissues remains a key limitation. Magnetic resonance imaging (MRI) has been successfully used to study many different plant models, including the study of in vivo changes during freezing. However, despite its benefits and past successes, the use of MRI in plant sciences remains low, likely due to limited access, high costs, and associated engineering challenges, such as keeping samples frozen for cold hardiness studies. To address this latter need, a novel device for keeping plant specimens at freezing temperatures during MRI is described. RESULTS: The device consists of commercial and custom parts. All custom parts were 3D printed and made available as open source to increase accessibility to research groups who wish to reproduce or iterate on this work. Calibration tests documented that, upon temperature equilibration for a given experimental temperature, conditions between the circulating coolant bath and inside the device seated within the bore of the magnet varied by less than 0.1 °C. The device was tested on plant material by imaging buds from Vaccinium macrocarpon in a small animal MRI system, at four temperatures, 20 °C, - 7 °C, - 14 °C, and - 21 °C. Results were compared to those obtained by independent controlled freezing test (CFT) evaluations. Non-damaging freezing events in inner bud structures were detected from the imaging data collected using this device, phenomena that are undetectable using CFT. CONCLUSIONS: The use of this novel cooling and freezing device in conjunction with MRI facilitated the detection of freezing events in intact plant tissues through the observation of the presence and absence of water in liquid state. The device represents an important addition to plant imaging tools currently available to researchers. Furthermore, its open-source and customizable design ensures that it will be accessible to a wide range of researchers and applications.

Vernalization shapes shoot architecture and ensures the maintenance of dormant buds in the perennial Arabis alpina.

Perennials have a complex shoot architecture with axillary meristems organized in zones of differential bud activity and fate. This includes zones of buds maintained dormant for multiple seasons and used as reservoirs for potential growth in case of damage. The shoot of Arabis alpina, a perennial relative of Arabidopsis thaliana, consists of a zone of dormant buds placed between subapical vegetative and basal flowering branches. This shoot architecture is shaped after exposure to prolonged cold, required for flowering. To understand how vernalization ensures the maintenance of dormant buds, we performed physiological and transcriptome studies, followed the spatiotemporal changes of auxin, and generated transgenic plants. Our results demonstrate that the complex shoot architecture in A. alpina is shaped by its flowering behavior, specifically the initiation of inflorescences during cold treatment and rapid flowering after subsequent exposure to growth-promoting conditions. Dormant buds are already formed before cold treatment. However, dormancy in these buds is enhanced during, and stably maintained after, vernalization by a BRC1-dependent mechanism. Post-vernalization, stable maintenance of dormant buds is correlated with increased auxin response, transport, and endogenous indole-3-acetic acid levels in the stem. Here, we provide a functional link between flowering and the maintenance of dormant buds in perennials.

Studies on germination of dormant bud and seed of Elaeagnus mollis under tisssue culturing / 中草药

Object To study the factors causing the dormancy of seed and to broaden the scope in the choice of explant for the tissue culture of the rare and near extinct plant Elaeagnus mollis Diels.Methods Dormant buds from one-year old branch collected each month from Autumn to next Spring, and seed of the very year were selected and cultured on basic media with the addition of 6-BA, NAA and GA 3 Results The germination rate of dormant bud follows the regular order of the months when it was taken. Those taken in Autumn declined every month until next Spring, and the nrevived monthly with those taken in March showing the most prosperous germination.The dormant bud can germinate into shoots. When seed was cultured, there are several factors leading to its dormancy: hardness of the mesocarp, keratinization of periderm, refractory to water permeation and the presence of certain inhibitory subtances. Seed kernal is rich in nutrition, but susceptible to bacterial infection leading to its decay. But the germination rate was well over 70% which can be cultured to give bacteria free shoots. Conclusion Both dormant buds obtained in March and bacteria free shoots germinated from naked kernal can be used as new sources for the tissue culture of E mollis .