The transcription factor MYB1 activates DGAT2 transcription to promote triacylglycerol accumulation in sacha inchi (Plukenetia volubilis L.) leaves under heat stress.

Triacylglycerol (TAG) accumulation is frequently triggered in vegetative tissues experiencing heat stress, which may increases plant basal plant thermo-tolerance by sequestering the toxic lipid intermediates that contribute to membrane damage or cell death under stress conditions. However, stress-responsive TAG biosynthesis and the underlying regulatory mechanisms are not fully understood. Here, we investigated the lipidomic and transcriptomic landscape under heat stress in the leaves of sacha inchi (Plukenetia volubilis L.), an important oilseed crop in tropical regions. Under heat stress (45 °C), the content of polyunsaturated TAGs (e.g., TAG18:2 and TAG18:3) and total TAGs were significantly higher, while those of unsaturated sterol esters, including ZyE 28:4, SiE 18:2 and SiE 18:3, were dramatically lower. Transcriptome analysis showed that the expression of PvDGAT2-2, encoding a type II diacylglycerol acyltransferase (DGAT) that is critical for TAG biosynthesis, was substantially induced under heat stress. We confirmed the function of PvDGAT2-2 in TAG production by complementing a yeast mutant defective in TAG biosynthesis. Importantly, we also identified the heat-induced transcription factor PvMYB1 as an upstream activator of PvDGAT2-2 transcription. Our findings on the molecular mechanism leading to TAG biosynthesis in leaves exposed to heat stress have implications for improving the biotechnological production of TAGs in vegetative tissues, offering an alternative to seeds.

Storage of orchid pollinia with varying lipid thermal fingerprints.

Orchid pollinia have the potential to make a valuable contribution to current techniques of germplasm storage and assisted reproduction, yet information regarding their preservation and their ability to remain viable over time is currently limited. Dactylorhiza fuchsii and Disa uniflora were used as models for investigating potential techniques for storing orchid pollinia. Initially, freshly harvested pollinia of Dact. fuchsii were incubated at 25 °C and 100% RH (relative humidity) for up to 7 days and germinated in vitro. For pollinia from both species, moisture sorption isotherms were constructed and thermal fingerprints generated using differential scanning calorimetry (DSC). Pollinia were stored at three temperatures (5, - 18 and - 196 °C) after equilibration at four different RHs (5, 33, 50 and 75%) and germinated. The isotherms and DSC results varied between species. Compared with D. uniflora, pollinia of Dact. fuchsii consistently equilibrated at higher moisture content (MC) for each RH, had less detectable lipids by DSC and had shorter lifespans, remaining viable after 3-4 months only at - 20 and - 196 °C and at low RH (5 and 33%). Both species' pollinia stored well at - 20 °C and - 196 °C, although there was some evidence of a small loss of viability under cryopreservation. In conclusion, pollen of these two species can be stored successfully for at least 3-4 months, and to maximize the pre-storage quality, it is recommended that fresh pollen is collected from flowers just prior to anthesis.

Frozen beauty: The cryobiotechnology of orchid diversity.

Orchids (Orchidaceae) are one of the most diverse plant groups on the planet with over 25,000 species. For over a century, scientists and horticulturalists have been fascinated by their complex floral morphology, pollinator specificity and multiple ethnobotanical uses, including as food, flavourings, medicines, ornaments, and perfumes. These important traits have stimulated world-wide collection of orchid species, often for the commercial production of hybrids and leading to frequent overexploitation. Increasing human activities and global environmental changes are also accelerating the threat of orchid extinction in their natural habitats. In order to improve gene conservation strategies for these unique species, innovative developments of cryopreservation methodologies are urgently needed based on an appreciation of low temperature (cryo) stress tolerance, the stimulation of recovery growth of plant tissues in vitro and on the 'omics' characterization of the targeted cell system (biotechnology). The successful development and application of such cryobiotechnology now extends to nearly 100 species and commercial hybrids of orchids, underpinning future breeding and species conservation programmes. In this contribution, we provide an overview of the progress in cryobanking of a range of orchid tissues, including seeds, pollen, protocorms, protocorm-like bodies, apices excised from in vitro plants, cell suspensions, rhizomes and orchid fungal symbionts. We also highlight future research needs.

Desiccation tolerance, longevity and seed-siring ability of entomophilous pollen from UK native orchid species.

BACKGROUND AND AIMS: Pollinator-limited seed-set in some terrestrial orchids is compensated for by the presence of long-lived flowers. This study tests the hypothesis that pollen from these insect-pollinated orchids should be desiccation tolerant and relatively long lived using four closely related UK terrestrial species; Anacamptis morio, Dactylorhiza fuchsii, D. maculata and Orchis mascula. METHODS: Pollen from the four species was harvested from inflorescences and germinated in vitro, both immediately and also after drying to simulate interflower transit. Their tolerance to desiccation and short-term survival was additionally assessed after 3 d equilibration at a range of relative humidities (RHs), and related to constructed sorption isotherms (RH vs. moisture content, MC). Ageing of D. fuchsii pollen was further tested over 2 months against temperature and RH, and the resultant survival curves were subjected to probit analysis, and the distribution of pollen death in time (σ) was determined. The viability and siring ability, following artificial pollinations, were determined in D. fuchsii pollen following storage for 6 years at -20 °C. KEY RESULTS: The pollen from all four species exhibited systematic increases in germinability and desiccation tolerance as anthesis approached, and pollen from open flowers generally retained high germinability. Short-term storage revealed sensitivity to low RH, whilst optimum survival occurred at comparable RHs in all species. Similarly, estimated pollen life spans (σ) at differing temperatures were longest under the dry conditions. Despite a reduction in germination and seeds per capsule, long-term storage of D. fuchsii pollen did not impact on subsequent seed germination in vitro. CONCLUSIONS: Substantial pollen desiccation tolerance and life span of the four entomophilous orchids reflects a resilient survival strategy in response to unpredictable pollinator visitation, and presents an alternative approach to germplasm conservation.

Post desiccation germination of mature seeds of tea (Camellia sinensis L.) can be enhanced by pro-oxidant treatment, but partial desiccation tolerance does not ensure survival at -20°C.

The maximal potential desiccation tolerance (MPDT) of tea (Camellia sinensis) seeds has been a matter of debate for decades. Here we assessed the ability of tea seeds from three sites in China to germinate after desiccation. Desiccation tolerance was greatest in Kunming, followed by Puer and Lincang, with Kunming seeds tolerating drying to 8% moisture content (MC), or ∼0.5 water activity (a(w)). Such tolerance was observed in Lincang seeds only when hydrogen peroxide (H2O2) at 0.5 or 1M was applied to seeds, indicating a stimulatory role for H2O2 in post-desiccation germination. Puer seeds exhibited MPDT of 16% MC (∼0.7 a(w)). Therefore, seeds from all three sites were not recalcitrant. The length of the dry season after dispersal and the high ratio of seed coat to seed mass (>0.3) support the observation of non-recalcitrant behaviour. The seeds were not immature, as the lipid signal in embryonic axes mirrored that of the cotyledons (30% oil). Even after high survival [>60% total germination (TG)] on drying to 10-13% MC, no Kunming seeds tolerated 1 month storage at -20 °C coinciding with lipid transitional changes at this temperature. The results indicate that tea seeds from China are neither recalcitrant nor storable at -20 °C.

Quantification of seed oil from species with varying oil content using supercritical fluid extraction.

INTRODUCTION: The quantity and composition of seed oil affects seed viability and storability and hence the value of a species as a resource for nutrition and plant conservation. Supercritical fluid extraction with carbon dioxide (SFE-CO2) offers a rapid, environmentally friendly alternative to traditional solvent extraction. OBJECTIVE: To develop a method using SFE-CO2 to quantify the seed oil content in a broad range of species with high to low oil contents. METHODOLOGY: Seed oil was extracted using SFE-CO2 from four crop species representing high, medium and low oil content: Helianthus annuus, Asteraceae, with ca. 55% oil; Brassica napus, Brassicaceae, with ca. 50% oil; Glycine max, Fabaceae, with ca. 20% oil; and Pisum sativum, Fabaceae, with ca. 2% oil. Extraction pressures of 5000, 6000 and 7500 psi and temperatures of 40, 60 and 80 degrees C were examined and a second step using 15% ethanol as a modifier included. Oil yields were compared with that achieved from Smalley Butt extraction. The optimised SFE-CO2 method was validated on six species from taxonomically distant families and with varying oil contents: Swietenia humilis (Meliaceae), Stenocereus thurberi (Cactaceae), Sinapis alba (Brassicaceae), Robinia pseudoacacia (Fabaceae), Poa pratensis (Poaceae) and Trachycarpus fortunei (Arecaceae). RESULTS: The two-step extraction at 6000 psi and 80 degrees C produced oil yields equivalent to or higher than Smalley Butt extraction for all species, including challenging species from the Brassicaceae family. CONCLUSION: SFE-CO2 enables the rapid analysis of seed oils across a broad range of seed oil contents.