Dormant Vegetative Buds Research Articles

Genome-Wide Identification and Expression Analysis of SnRK2 Gene Family in Dormant Vegetative Buds of Liriodendron chinense in Response to Abscisic Acid, Chilling, and Photoperiod.

Protein kinases play an essential role in plants’ responses to environmental stress signals. SnRK2 (sucrose non-fermenting 1-related protein kinase 2) is a plant-specific protein kinase that plays a crucial role in abscisic acid and abiotic stress responses in some model plant species. In apple, corn, rice, pepper, grapevine, Arabidopsis thaliana, potato, and tomato, a genome-wide study of the SnRK2 protein family was performed earlier. The genome-wide comprehensive investigation was first revealed to categorize the SnRK2 genes in the Liriodendron chinense (L. chinense). The five SnRK2 genes found in the L. chinense genome were highlighted in this study. The structural gene variants, 3D structure, chromosomal distributions, motif analysis, phylogeny, subcellular localization, cis-regulatory elements, expression profiles in dormant buds, and photoperiod and chilling responses were all investigated in this research. The five SnRK2 genes from L. chinense were grouped into groups (I–IV) based on phylogeny analysis, with three being closely related to other species. Five hormones-, six stress-, two growths and biological process-, and two metabolic-related responsive elements were discovered by studying the cis-elements in the promoters. According to the expression analyses, all five genes were up- and down-regulated in response to abscisic acid (ABA), photoperiod, chilling, and chilling, as well as photoperiod treatments. Our findings gave insight into the SnRK2 family genes in L. chinense and opened up new study options.

An Early Calcium Loading during Cherry Tree Dormancy Improves Fruit Quality Features at Harvest

The possible role of an early calcium application via sprays (0.25, 0.5 and 1M CaCl2) on dormant buds to improve sweet cherry (cv. Ferrovia) fruit quality at harvest was investigated. Fruit quality characteristics were also investigated in response to the age of spurs, the ripening stage, and their interactions. Results indicate that calcium enters the dormant flower buds and the phloem but not to the dormant vegetative buds. At harvest, the levels of Zn, Mn, and Cu were declined in fruits by increasing CaCl2 doses of sprays. Fruit respiratory activity was higher and on–tree fruit cracking was lower in red-colour (unripe) cherries as well as in fruit that was produced by 2-year-old short spurs or by Ca-treated buds. Differences in the sweet cherry skin metabolic profiles were identified. Fruit produced from Ca-exposed spurs exhibited lower levels of ribose and other cell-wall-related sugars and higher sucrose, maltose, and quininic acid levels. Nutrient shift was increased in red cherries, while anthocyanins were boosted in the black ones. PCA analysis was performed between the high dose of calcium spray and a control for mineral element content and cherry quality traits. This study illustrates that the high dose of calcium application during bud dormancy can effectively improve sweet cherry fruit characteristics, in terms of calcium content, cracking incidence, and fruit set. Overall, the present study contributes to a better understanding of the impact of calcium nutrition in fruit crops, which will provide references for alternative nutrient management and quality control in sweet cherry production.

Influence of light, water stress and shrub cover on sapling survival and height growth: the case of A. unedo, F. ornus and S. domestica under Mediterranean climate

Resprouter species are of great interest for Mediterranean forest resilience as they can survive fire or drought by activating dormant vegetative buds to produce regrowth. Understanding which factors control the early growth and survival of these species is needed to develop strategies to enhance stand resilience. In this study, three broadleaved resprouter species, two trees—Fraxinus ornus and Sorbus domestica—and a shrub—Arbutus unedo—were planted under a gradient of pine cover. Sapling survival and height growth were monitored for 6 years, 2 years after their plantation, as well as light and shrub cover. We developed a two-strata forest water balance model, which produces a water stress index. We aimed to determine the effect of light, water stress and shrub cover on sapling survival and height growth using a modelling approach. We found that high levels of transmittance had a negative impact on both survival and height growth of F. ornus and S. domestica, while it had a positive effect on A. unedo growth. Water stress was found to negatively affect survival and height growth of all species. Lastly, shrub cover had an overall positive effect on the survival and growth of saplings, indicating a facilitating effect of the shrubs. Our results suggest that shade-tolerant tree species like F. ornus and S. domestica are more suited to moderate and light cover, while photoinhibition-tolerant shrub species like A. unedo are more adapted to open conditions. To maximize sapling survival and growth, we recommend preserving the shrubby understorey when present.

Genotypic variation in Norway spruce correlates to fungal communities in vegetative buds.

The taxonomically diverse phyllosphere fungi inhabit leaves of plants. Thus, apart from the fungi's dispersal capacities and environmental factors, the assembly of the phyllosphere community associated with a given host plant depends on factors encoded by the host's genome. The host genetic factors and their influence on the assembly of phyllosphere communities under natural conditions are poorly understood, especially in trees. Recent work indicates that Norway spruce (Picea abies) vegetative buds harbour active fungal communities, but these are hitherto largely uncharacterized. This study combines internal transcribed spacer sequencing of the fungal communities associated with dormant vegetative buds with a genome‐wide association study (GWAS) in 478 unrelated Norway spruce trees. The aim was to detect host loci associated with variation in the fungal communities across the population, and to identify loci correlating with the presence of specific, latent, pathogens. The fungal communities were dominated by known Norway spruce phyllosphere endophytes and pathogens. We identified six quantitative trait loci (QTLs) associated with the relative abundance of the dominating taxa (i.e., top 1% most abundant taxa). Three additional QTLs associated with colonization by the spruce needle cast pathogen Lirula macrospora or the cherry spruce rust (Thekopsora areolata) in asymptomatic tissues were detected. The identification of the nine QTLs shows that the genetic variation in Norway spruce influences the fungal community in dormant buds and that mechanisms underlying the assembly of the communities and the colonization of latent pathogens in trees may be uncovered by combining molecular identification of fungi with GWAS.

Dormancy release and flowering time in Ziziphus jujuba Mill., a “direct flowering” fruit tree, has a facultative requirement for chilling

In deciduous fruit trees, the effect of chilling on flowering has mostly been investigated in the “indirect flowering” group, characterized by a period of rest between flower bud formation and blooming. In the present study, we explored the effects of chilling and chilling deprivation on the flowering of Ziziphus jujuba, a temperate deciduous fruit tree belonging to the “direct flowering” group, in which flower bud differentiation, blooming and fruit development occur after dormancy release, during a single growing season. Dormancy release, vegetative growth and flowering time in Z. jujuba cv. Ben-Li were assessed following several treatments of chilling. Chilling treatments quantitatively decreased the timing of vegetative bud dormancy release, thereby accelerating flowering, but had no effect on the time from dormancy release to flowering. Trees grown at a constant temperature of 25°C, without chilling, broke dormancy and flowered, indicating the facultative character of chilling in this species. We measured the expression of Z. jujuba LFY and AP1 homologues (ZjLFY and ZjAP1). Chilling decreased ZjLFY expression in dormant vegetative buds but had no effect on ZjAP1expression, which reached peak expression before dormancy release and at anthesis.In conclusion, chilling is not obligatory for dormancy release of Z. jujuba cv. Ben-Li vegetative buds. However, the exposure to chilling during dormancy does accelerate vegetative bud dormancy release and flowering.

Custom Microarray Analysis for Transcript Profiling of Dormant Vegetative Buds of Japanese Apricot during Prolonged Chilling Exposure

Bud dormancy is a critical developmental process for perennial plant survival, and also an important physiological phase that affects the next season’s growth of temperate fruit trees. Bud dormancy is regulated by multiple genetic factors, and affected by various environmental factors, tree age and vigor. To understand the molecular mechanism of bud dormancy in Japanese apricot (Prunus mume Sieb. et Zucc.), we constructed a custom oligo DNA microarray covering the Japanese apricot dormant bud ESTs referring to the peach (P. persica) genome sequence. Because endodormancy release is a chilling temperature-dependent physiological event, genes showing chilling-mediated differential expression patterns are candidates to control endodormancy release. Using the microarray constructed in this study, we monitored gene expression changes of dormant vegetative buds of Japanese apricot during prolonged artificial chilling exposure. In addition, we analyzed seasonal gene expression changes. Among the 58539 different unigene probes, 2345 and 1059 genes were identified as being more than twofold up-regulated and down-regulated, respectively, following chilling exposure for 60 days (P < 0.05). Cluster analysis suggested that the expression of the genes showing expression changes by artificial chilling exposure were coordinately regulated by seasonal changes. The down-regulated genes included P. mume DORMANCY-ASSOCIATED MADS-box genes, which supported previous quantitative RT-PCR and EST analyses showing that these genes are repressed by prolonged chilling exposure. The genes encoding lipoxygenase were markedly up-regulated by prolonged chilling. Our parametric analysis of gene-set enrichment suggested that genes related to jasmonic acid (JA) and oxylipin biosynthesis and metabolic processes were significantly up-regulated by prolonged chilling, whereas genes related to circadian rhythm were significantly down-regulated. The results obtained from microarray analyses were verified by quantitative RT-PCR analysis of selected genes. Taken together, we have concluded that the microarray platform constructed in this study is applicable for deeper understanding of the molecular network related to agronomically important bud physiology, including dormancy release.

12. From the field to the tank: The cryo-banking of ancient Italian apple germplasm by the dormant-bud technique

The cryopreservation of dormant vegetative buds, proposed for the first time by Sakai in early 1960s for winter-collected twigs of poplar and willow, was afterwards adapted by Forsline and co-workers to the conservation in liquid nitrogen of uni-nodal shoot segments of apple cultivars and used by the National Plant Germplasm System (NPGS), in the USA, to back-up in-field apple germplasm. In short, the dormant-bud protocol is based on winter collection of naturally cold-hardened scions from which 35 mm uni-nodal segments are cut, desiccated, slow cooled (−1 °C/h) down to −30 °C, and stored in liquid nitrogen. In late spring, thawed and rehydrated segments are transferred to the greenhouse in a cold box where the individual buds are removed from the segments and used for vegetative propagation by chip budding onto rootstocks. Cryopreservation by the dormant-bud technique has the advantage of allowing the direct transfer of accessions from the field to the tank and vice versa , i.e., with no passage in tissue culture, which is necessary with the PVS2-based vitrification technique before and after storage in liquid nitrogen. For this reason, the dormant-bud cryopreservation of apple takes about 40% of the time and 50% of the labour necessary with the shoot-tip vitrification approach. Recently, the technique has moved from the USA to Europe, where programs for the adaptation of protocols to different winter climatic conditions are in progress in Italy, Denmark, Germany and Switzerland. Italy, for instance, is the highest producer of apple in the EU, with a market of over 2,200,000 tons of fruits in the year 2011. However, only three cultivars (‘Golden Delicious’, ‘Gala’ and ‘Red Delicious’) contribute for 70% of this wide-scale commercial production. Hence, Italy, as well as other European countries, is facing loss of diversity in ancient apple fruit germplasm. Because of that, in 2000, Veneto Agricoltura, the agricultural agency of the Veneto region (Italy), started a program aimed at conserving ancient apple cultivars, before they were lost forever. This program is focused on the documentation of specimen trees from old cultivars and their propagation and maintenance in in-field collection. The clonal orchard contains today about 200 cultivars. In order to guarantee a safe duplication of this valuable germplasm, cryopreservation by the dormant-bud technique was recently included in the programme, in collaboration with the CNR-IVALSA Institute of Florence. Up to now, an effective cryopreservation protocol has been developed for 19 accessions, with up to 100% of plants regrown after they were chip-budded with buds recovered from the storage in liquid nitrogen. Additional cultivars are introduced yearly in experimentation, with a careful evaluation for each one of the time required to desiccate uni-nodal segments below 30% of moisture content. A specific experiment is currently in progress aimed at evaluating the possibility to bypass one or more of the several steps of the original procedure. Moreover, in order to speed up the development and optimization of cryo-protocols for all the accessions, two post-conservation pre-grafting tests (i.e., the TTC and the electrolyte leakage assays) are under evaluation, with the aim to investigate bud and cambium cell survival to liquid nitrogen, as well as the possible correlation between results from these tests and from plant regrowth after grafting with cryopreserved buds.

Microsatellite DNA somaclonal variation in micropropagated trembling aspen (Populus tremuloides)

Microsatellite DNA markers of ten simple sequence repeat (SSR) loci were used to examine somaclonal variation in randomly selected micropropagated plantlets derived from three different Populus tremuloides donor trees (genotypes). The plantlets were obtained from tissue cultures of dormant vegetative buds, and those derived from the same donor tree, grown in the greenhouse, did not exhibit any sign of visible morphological variation. No microsatellite DNA variation was observed among 13 somaclones of one tree and 4 somaclones of another tree at eight of the ten SSR loci. However, despite the small number of micropropagated progeny per tree sampled, microsatellite DNA variation was detected among the plantlets derived from the same donor trees at two SSR loci. The primer pair for the SSR locus PTR5 revealed somaclonal variation in 1 out of the 13 plantlets obtained from one genotype, while the primer pair for the PTR2 SSR locus revealed somaclonal variation in one out of the four plantlets obtained from another genotype. The variation at the PTR2 locus resulted in the appearance of a new allele of increased size, possibly due to an addition of the repeat units, while the variation at the PTR5 locus resulted in the appearance of third allele, presumably due to the presence of a single extra chromosome or duplication of a chromosomal segment. These results demonstrate that the genetic fidelity of micropropagated plants of P. tremuloides cannot always be assured and somaclonal variation can occur even when tissues of well organized vegetative buds are used for tissue cultures; that somaclonal variation cannot always be detected at the gross morphological level; and that microsatellite DNA markers provide useful and sensitive markers for determining the clonal fidelity and somaclonal variation in P. tremuloides.

Preformation, architectural complexity, and developmental flexibility in Acomastylis rossii (Rosaceae)

The duration of preformation and the seasonal pattern of development were studied in the architecturally complex alpine perennial Acomastylis rossii. Each leaf and inflorescence requires 3 yr to progress from initiation through structural and functional maturity to senescence. As a consequence, three cohorts of preformed organs, initiated in successive years, are borne simultaneously by each individual plant. The oldest cohort matures immediately following snowmelt, after which no additional leaves are matured until the following spring. A second cohort remains below ground in the apical bud and continues development, while a third cohort is initiated. Initiation and development of primordia proceed below ground throughout the summer and continue for at least 2.5 mo after aboveground structures have senesced. Acomastylis rossii maintains numerous dormant vegetative buds containing preformed leaf primordia in the axils of senesced leaves. Developmental preformation has been widely reported in arctic and alpine tundra environments and has been theorized to severely constrain rapid responses to environmental variation. The presence of many such preformed structures may mitigate some of the constraint on plant response to environmental variation imposed by the long developmental trajectories of leaves and inflorescences in apical buds.

Cryopreservation of Cold-tender Apple Germplasm

Unlike cold-hardy apple germplasm, dormant vegetative buds from cold-tender accessions require stabilization of meristematic tissue to protect against injury during desiccation and cryopreservation. Dormant buds of six apple cultivars [Malus sylvestris (L.) Mill. var. domestica (Borkh.) Mansf. `Cox's Orange Pippin', `Einshemer', `Golden Delicious', `Jonagold', `K-14', and `Mutsu'] collected at specific intervals in 1993, 1994, and 1995 at Geneva, N.Y., were stabilized by encapsulation in 5% alginate, treated with step-wise imbibition of 0.5 to 1.0 m sucrose and 0.2 m raffinose solution, and desiccated with forced air at 0 °C. Sugar-alginate stabilization reduced injury during desiccation, increased cold-hardiness of the six cold-tender cultivars frozen to -30 °C, and improved recovery following cryopreservation of buds collected before optimal cold acclimation was attained. Sucrose tissue levels did not increase following stabilization treatment, but levels of glucose and fructose, and of an unknown disaccharide increased. This procedure used nontoxic cryoprotectants, and has potential to expand the scope of dormant bud cryopreservation to include cold-tender apple germplasm.

Tissue culture responses of explants taken from branch sources with different degrees of juvenility in mature black locust (Robinia pseudoacacia) trees.

We studied the in vitro responses of cambial tissue and dormant vegetative buds obtained from top and epicormic branches of three mature black locust (Robinia pseudoacacia L.) trees. Cambial tissues isolated from epicormic branches produced more callus than cambial tissues isolated from top branches, whereas in vitro shoot cultures derived from buds excised from top branches grew faster than those derived from buds excised from epicormic branches. There were no significant differences between the two branch sources in in vitro bud break or shoot multiplication from bud explants or cambial-derived callus tissue, respectively. Furthermore, the top branches, generally considered to be the most mature in a tree, were not recalcitrant in terms of morphogenic capacity compared to epicormic branches.

Cryopreservation of Dormant Vegetative Buds from Sour Cherry

The dormant vegetative bud method for cryopreservation has been successfully applied to many lines of apple. We examined this method for five cultivars (Kentish, Montmorency, Meteor, North Star, Schatten Morelle) of sour cherry (Prunus cerasus L.) with the aim of developing long-term storage at NSSL. Singlebud nodal sections (35 cm) were desiccated to 25%, 30%, or 35% moisture before cooling at 1°C/hour to –30°C and holding for 24 hours. Sections were then directly placed in storage in the vapor phase above liquid nitrogen (about – 160°C). Warmed samples were rehydrated and patch budded at Geneva to assess viability. Sections that were either undried, dried but unfrozen, or dried and cooled to –30°C survived very well. For samples then cooled to –160°C, highest viabilities for each line occurred with the 25% moisture level, although fairly high viabilities also were observed at 30% and 35% moistures. Cryopreserved buds from four lines directly developed into a single shoot; buds from Montmorency formed a shoot from a lateral within the bud, suggesting that the terminal meristem died but that axillary meristems within the bud survived and formed a shoot or multiple shoots. Nineteen lines were harvested in January 1996 for long term storage of sour cherry germplasm under cryogenic conditions.

Seasonal and geographic origin effects on cold hardiness of white spruce buds, foliage, and stems

Cold hardiness of dormant vegetative buds from white spruce (Piceaglauca (Moench) Voss) trees originating from throughout the species' North American range was measured by differential thermal analysis between August and April at Vernon, B.C. Buds were least hardy in early fall (low temperature exotherm −12 to −25 °C) and most hardy midwinter (low temperature exotherms −20 to −34 °C). Buds of northerly origin trees were more hardy in early fall (September–October) than buds of more southerly origin trees. Similar results were observed for foliage and stem tissue in early fall using electrolyte leakage measurements following −16 °C freezing.

618 PB 322 CRYOPRESERVATION OF COLD-TENDER APPLE, Malus domestica

Cryopreservation of mature dormant vegetative buds is a useful method to preserve germplasm of a large number of cold hardy apple cultivars. However, cold tender cultivars have proven to be much more difficult to cryopreserve. Eight cultivars were harvested in September 1993 at Geneva, NY before developing cold hardiness naturally. The twigs were encapsulated with 5% alginate and treated with stepwise imbibition of 0.5 to 1.0 M sucrose. The samples were desiccated over glycerol at 0C. Half of the samples were plunged directly into liquid nitrogen (IN) and the other half were first cooled slowly to -30C. The twigs that had been exposed to prefreezing conditions showed the highest survival (20 to 100%). The samples that were plunged directly in LN survived poorly (0 to 20 %). Samples without encapsulation and no sucrose imbibition had 0% survival. We conclude that this protocol opens up the possibility to expand cryopreservation of cold tender apple cultivars, presently grown only in field gene banks, at great expense and inconvenience.

A plant gene with homology to D-myo-inositol-3-phosphate synthase is rapidly and spatially up-regulated during an abscisic-acid-induced morphogenic response in Spirodela polyrrhiza.

The induction of turions (dormant vegetative buds) by the plant growth regulator abscisic acid (ABA) in the aquatic angiosperm Spirodela polyrrhiza is one of the few examples of a plant hormone triggering a morphogenic response. In order to aid our understanding of the molecular basis of this phenomenon a cDNA library has been prepared from ABA-induced S. polyrrhiza and by differential screening a number of early ABA-up-regulated genes have been isolated. This paper reports the characterization of one such cDNA (tur1) from S. polyrrhiza which codes for a protein highly homologous to yeast D-myo-inositol-3-phosphate synthase (EC.5.5.1.4), the catalyst for the first committed step in inositol biosynthesis. This is the first reported cloning of a plant gene involved in inositol production. It is shown by Northern analysis and in situ hybridization that ABA induces a rapid preferential up-regulation of this gene and that this is localized to the stolon tissue. The stolon connects the developing turion to the node of the mother frond (the meristematic region involved in the production of turions which arise as a morphogenic response to this hormone). This increase in tur1 transcript level is accompanied by a rise in the activity of D-myo-inositol-3-phosphate synthase. The possible role of inositol in ABA-induced events during turion formation is discussed.

Ecology of a forest weed of Newfoundland: vegetative regeneration strategy of Kalmia angustifolia

The vegetative regeneration characteristics of Kalmia angustifolia were studied by examining the near ground and below-ground structures of the plant. Three methods of vegetative regeneration were observed in Kalmia: layering, stem-base sprouting, and rhizomatous growth, which make the plant more resilient than other ericaceous plants of similar stature following disturbance. Layering occurred only in tall old Kalmia of undisturbed habitats. Stem-base sprouting and rhizomatous growth were found to be the most common and most effective modes of vegetative regeneration. These methods were common in both disturbed and undisturbed habitats. Three types of rhizome systems, namely primary, secondary, and tertiary rhizomes, were identified and described for the first time in Kalmia. Secondary rhizomes were found to be the most effective organ for vegetative spread of the plant. Some secondary rhizomes may grow more than 1 m in one growing season. A 5-year-old Kalmia may produce, on average, 36 secondary rhizomes with a total length of 750 cm. Secondary rhizomes of Kalmia first grow horizontally, then come aboveground growing vertically and exhibiting apical dominance. Large numbers of dormant vegetative buds (three at each node) and adventitious roots are produced at the rhizome nodes. Transverse sections of current-year rhizome showed many trichomes. The cells of the pith are thick-walled and are characterized by large lysigenous cavities. The anatomy of axillary buds and leaf primordia of the aboveground stems of Kalmia were similar to buds and bud scales of the rhizome. Key words: Kalmia, disturbance, fire, regeneration strategy, rhizome, trichomes.

DEHYDRATION OF DORMANT APPLE BUDS AT DIFFERENT STAGES OF COLD ACCLIMATION TO INDUCE CRYOPRESERVABILITY IN DIFFERENT CULTIVARS

Survival in liquid nitrogen of dormant vegetative buds from several cold-hardened apple cultivars was greater with buds which were dehydrated prior to cryopreservation than with nondehydrated buds. Buds collected early in the cold-acclimating period suffered injury as a result of dehydration, but the percent survival of the dehydrated buds, after storage in liquid nitrogen, was greater than that of nondehydrated buds. As cold acclimation progressed, buds became more resistant to the dehydration stress and survival in liquid nitrogen increased following dehydration for all cultivars. Survival in liquid nitrogen of nondehydrated buds increased for LN2-hardy but not for LN2-tender cultivars, as cold acclimation progressed. The magnitude of the dehydration-induced increase in survival in liquid nitrogen was cultivar dependent.Key words: Malus domestica, apple, cryopreservation, gene resources, cold hardiness