Seed maturation and effect of temperature regime on Trema micrantha (L.) Blume seed germination

T. micrantha (L.) Blume (Ulmaceae), a common pioneer tree species in Brazil, is used in the restoration of degraded areas. The fruits are fleshy and indehiscent, with only one water impermeable seed. During the fruiting period, fruits of different colours are found at the same time on the same branch. This research aimed to correlate fruit colour with other physical indicators of seed maturity and to Verify the effect of temperature regime on seed germination. Collected fruits were separated in to green, green-red and red colour and for each of these maturation stages, size, moisture content and dry matter of both fruits and seeds were determined. Seeds were scarified with sulphuric acid and submitted to a germination test conducted at constant (20 degreesC, 30 degreesC and 40 degreesC) and alternating (20-30 degreesC, 30-40 degreesC and 20-40 degreesC) temperatures for 15 weeks. Seed germination percentage and speed were analysed after five weeks and the fmal percentage of germinated and Viable seeds after 15 weeks. Seed maturity is attained when the fruits are green-red. At this stage, moisture content was about 64% for fruits and 10% for seeds. Alternating temperature was required for seed germination and 20-30 degreesC was the best option. Most seeds had germinated after five weeks, providing mature seeds, acid scarification and alternating temperature were used.

Concentrações de sacarose no desenvolvimento in vitro e na aclimatização de Cattleya loddigesii Lindley

This work aimed to study the influence of sucrose in the culture medium for in vitro growth and acclimatization of the epiphytic orchid Cattleya loddigesii. Five sucrose treatments (absence, 10, 20, 30 and 40g L-1) were used in a randomic experimental design. Mature seeds were sowed in 1/2 MS culture medium and after 90 days the plantlets (1.0 +/-0.2 cm) were inoculated between the treatments, whereby they were remained more 90 days. After 180 days of the beginning of the experiment the plantlets were removed from the flasks and evaluated the number of roots, shoot length, number of leafs, total dry weight and photosynthetic pigments. Survival percentage was evaluated after 75 days of acclimatization. The data of biometric variables were analyzed by Anovaand polynomial regression (p<0.05). Theothers data were submitted to the Anova and the means compared by the Tukey test (p<0.05). The sucrose concentration of 20g L-1 favored the in vitro growth in all evaluated parameters, showed higher production of chlorophyll a, total chlorophyll and carotenoids, in addition to increased survival under ex vitro condition. The sucrose concentration of 20g L-1 in the culture medium was the most efficient among the tested concentrations both for in vitro growth and ex vitro establishment of Cattleya loddigesii.

Cryopreservation, early seedling development, and genetic stability of Oncidium flexuosum Sims

An efficient cryopreservation protocol was developed for mature seeds of Oncidium flexuosum Sims. Seed morphology, protocorm formation, and early seedling development were also assessed. The effects of phloroglucinol and Supercool X-1000® as cryoprotectant additives in the vitrification solution were investigated. Dehydration using the plant vitrification solution 2 (PVS2) for 60 and 120 min prior to immersion in liquid nitrogen promoted the highest frequency of in vitro seed germination 6 weeks following culture on half-strength Murashige and Skoog (1/2 MS) medium. Mature seeds submitted to vitrification for 120 min in PVS2 and 1 % phloroglucinol at 0 °C enhanced germination by 68 %, whereas in PVS2 and 1 % Supercool X-1000® germination was just moderately enhanced (26 %). In vitro-germinating seedlings developed healthy shoots and roots without the use of plant growth regulators. After 6 months of growth, there were no differences between in vitro- and ex vitro-grown seedlings for various phenotypic characteristics, including shoot length, number of leaves, number and length of roots, and fresh and dry weight. Seedlings were transferred to greenhouse conditions and successfully acclimatized, further developing into normal plants with over 90 % survival. Comparative analysis of seedlings from control and vitrified seeds using flow cytometry indicated that no change in ploidy levels occurred as a result of cryopreservation, therefore maintaining seedlings genetic stability. In this study, vitrification with PVS2 for 120 min with the addition of 1 % phloroglucinol offers a simple, safe, and feasible protocol for cryopreservation of O. flexuosum mature seeds. © 2013 Springer Science+Business Media Dordrecht.

Mecanismos de controle de tolerância à dissecação em sementes de Caesalpinia echinata LAM. (Pau-Brasil) e Caesalpinia peltophoroides BENTH. (Sibipiruna)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Criopreservação, indução de poliploidia e avaliação da estabilidade genética de orquídeas

Pós-graduação em Agronomia (Genética e Melhoramento de Plantas) - FCAV

In vitro conservation of Dendrobium germplasm

Dendrobium is a large genus in the family Orchidaceae that exhibits vast diversity in floral characteristics, which is of considerable importance to orchid breeders, biotechnologists and collectors. Native species have high value as a result of their medicinal properties, while their hybrids are important as ornamental commodities, either as cut flowers or potted plants and are thus veritable industrial crops. Thus, preservation of Dendrobium germplasm is valuable for species conservation, breeding programs and the floriculture industry. Cryopreservation represents the only safe, efficient and cost-effective long-term storage option to facilitate the conservation of genetic resources of plant species. This review highlights 16 years of literature related to the preservation of Dendrobium germplasm and comprises the most comprehensive assessment of thorough studies performed to date, which shows reliable and reproducible results. Air-drying, encapsulation-dehydration, encapsulation-vitrification, vitrification and droplet-vitrification are the current cryopreservation methodologies that have been used to cryopreserve Dendrobium germplasm. Mature seeds, pollen, protoplasts, shoot primordia, protocorms and somatic embryos or protocorm-like bodies (PLBs) have been cryopreserved with different levels of success. Encapsulation-vitrification and encapsulation-dehydration are the most used protocol, while PLBs represent the main explant explored.

Regenerative morphological traits in a woody species community in Tumbesian tropical dry forest.

The study of functional morphological traits enables us to know fundamental aspects of the dynamics of plant communities in local and global habitats. Regenerative morphological traits play an important role in defining plant history and ecological behavior. Seed and fruit characteristics determine to a large extent the patterns for dispersal, germination, establishment and seedling recruitment a given species exhibits on its natural habitat. Despite their prominent role, seed and fruit traits have been poorly studied at the community level of woody plant species in neo-tropical dry forests. In the present study we aimed at i) evaluate the functional role of morphological traits of seeds, fruits and embryo in woody plant species; ii) determine which are the morphological patterns present in seeds collected from the community of woody species that occur in neo-tropical dry forests; and iii) compare woody plant species seed mass values comparatively between neo-tropical dry and tropical forests. To do so, mature seeds were collected from 79 plant species that occur in the Tumbesian forest of Southwest Ecuador. The studied species included the 42 and 37 most representative tree and shrubbery species of the Tumbesian forest respectively. A total of 18 morphological traits (seven quantitative and 11 qualitative) were measured and evaluated in the seeds, fruits and embryos of the selected species, and we compared the seeds mass with other forest types. Our results showed a huge heterogeneity among traits values in the studied species. Seed mass, volume and number were the traits that vary the most at the community level, i.e. seed length ranged from 1.3 to 39 mm, and seed width from 0.6 to 25 mm. Only six embryo types were found among the 79 plant species. In 40 % of the cases, fully developed inverted embryos with large and thick cotyledons to store considerable amount of nutrients were recorded. We concluded that highly variable and functionally complementary morphological traits occur among the studied woody plants of the dry Tumbesian forest. The latter favors a plethora of behavioral mechanisms to coexist among woody species of the dry forest in response to the environmental stress that is typical of arid areas.

Expression Pattern of the Carrot EP3 Endochitinase Genes in Suspension Cultures and in Developing Seeds1

Carrot (Daucus carota) extracellular protein 3 (EP3) class IV endochitinases were previously identified based on their ability to rescue somatic embryos of the temperature-sensitive cell line ts11. Whole-mount in situ hybridization revealed that a subset of the morphologically distinguishable cell types in embryogenic and nonembryogenic suspension cultures, including ts11, express EP3 genes. No expression was found in somatic embryos. In carrot plants EP3 genes are expressed in the inner integumentary cells of young fruits and in a specific subset of cells located in the middle of the endosperm of mature seeds. No expression was found in zygotic embryos. These results support the hypothesis that the EP3 endochitinase has a “nursing” function during zygotic embryogenesis and that this function can be mimicked by suspension cells during somatic embryogenesis.

Environmental and developmental signals modulate proline homeostasis: evidence for a negative transcriptional regulator.

In many plants, osmotic stress induces a rapid accumulation of proline through de novo synthesis from glutamate. This response is thought to play a pivotal role in osmotic stress tolerance [Kishor, P. B. K., Hong, Z., Miao, G.-H., Hu, C.-A. A. and Verma, D. P. S. (1995) Plant Physiol. 108, 1387-1394]. During recovery from osmotic stress, accumulated proline is rapidly oxidized to glutamate and the first step of this process is catalyzed by proline oxidase. We have isolated a full-length cDNA from Arabidopsis thaliana, At-POX, which maps to a single locus on chromosome 3 and that encodes a predicted polypeptide of 499 amino acids showing significant similarity with proline oxidase sequences from Drosophila and Saccharomyces cerevisiae (55.5% and 45.1%, respectively). The predicted location of the encoded polypeptide is the inner mitochondrial membrane. RNA gel blot analysis revealed that At-POX mRNA levels declined rapidly upon osmotic stress and this decline preceded proline accumulation. On the other hand, At-POX mRNA levels rapidly increased during recovery. Free proline, exogenously added to plants, was found to be an effective inducer of At-POX expression; indeed, At-POX was highly expressed in flowers and mature seeds where the proline level is higher relative to other organs of Arabidopsis. Our results indicate that stress- and developmentally derived signals interact to determine proline homeostasis in Arabidopsis.

Crystal structure of mature 2S albumin from Moringa oleifera seeds

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Anatomical studies of the flower, fruit and seeds of Myrceugenia rufa (Myrtaceae)

Myceugenia rufa is a rare and endemic species from the coast of central Chile. There are no published studies describing flower, fruit or seed anatomy. Forty-two accessions were collected from across the geographic range of the species. Reproductive structures were fixed, dehydrated, embedded in paraffin, sectioned and stained with Safranin O and Fast green. Anatomy of floral buds, mature flowers, fruits and seeds was described. Reproductive anatomy matches that of other Myrtaceae, such as presence of druses, internal phloem and schizogenous secretory cavities in buds, flowers, fruits and seeds. The anatomy and development of reproductive structures of M. rufa might enhance the understanding for future studies regarding natural reproduction and conservation programs.

Synthesis of Trilaurin by Developing Pisa Seeds (Actinodaphne hookeri)

The developing seeds of Actinodaphne hookeri were investigated to delineate their ability to synthesize large amounts of trilaurin. Until 88 days after flowering the embryos contained 71% neutral lipids (NL) and 29% phospholipids (PL) and both these components contained C-16:0, C-18:0, C-18:2, and C-18:3 as the major fatty acids (FA). At 102 days after flowering the seeds began to accumulate triacylglycerols (TAG) and to synthesize lauric acid (C-12:0). By 165 days after flowering, when the seeds were mature, they contained about 99% NL and 1% FL. At this stage the TAG contained exclusively C-12:0, while the PL consisted of long-chain fatty acids (LCFA) only. Leaf lipids in contrast did not contain any C-12:0. Experiments on [1-C-14]acetate incorporation into developing seed slices showed that at 88 days after flowering only 4% of the label was in TAG, 1% in diacylglycerols (DAG), and 87% in FL. One hundred two days after flowering seeds incorporated only 2% of the label into TAG, 30% into DAG, and 64% into FL. In contrast at 114 days after flowering 71% of the label was incorporated into TAG, 25% into DAG, and only 2% into FL. Analysis of labeled FA revealed that up to 102 days after flowering it was incorporated only into LCFA, whereas at 114 days after flowering it was incorporated exclusively into C-12:0. Furthermore, 67% of the label in PL at 114 days after flowering was found to be dilaurylglycerophosphate. Analysis of the label in DAG at this stage showed that it was essentially in dilaurin species. These observations indicate the induction of enzymes of Kennedy pathway for the specific synthesis of trilaurin at about 114 days after flowering, Homogenates of seeds (114 days after flowering) incubated with labeled FA in the presence of glycerol-3-phosphate and coenzymes A and ATP incorporated 84% of C-12:0 and 61% of C-14:0, but not C-16:0, C-18:2, and C-18:3, into TAG. In contrast the LCFA were incorporated preferentially into FL. It is concluded that, between 102 and 114 days after flowering, a switch occurs in A. hookeri for the synthesis of C-12:0 and trilaurin which is tissue specific. Since the seed synthesizes exclusively C-12:0 at 114 days after flowering onwards and incorporates specifically into TAG, this system appears to be ideal for identifying the enzymes responsible for medium-chain fatty acid as well as trilaurin synthesis and for exploiting them for genetic engineering. (C) 1994 Academic Press, Inc.

Onset of germinability, desiccation tolerance and hardseededness in developing seeds of Peltophorum pterocarpum (DC) K. Heyne (Caesalpinioideae)

In the hot and dry conditions in which seeds of the tree legume Peltophorum pterocarpum develop and mature in Vietnam, seed moisture content declined rapidly on the mother plant from 87% at 42 d after flowering (DAF) to 15% at 70 DAF. Dry weight of the pods attained a maximum value at about 42 DAF, but seed mass maturity (i.e. the end of the seed-filling phase) occurred at about 62 DAF, at which time seed moisture content was about 45-48%. The onset of the ability of freshly collected seeds to germinate (in 63-d tests at 28-34degreesC) occurred at 42 DAF, i.e. about 20 d before mass maturity. Full germination (98%) was attained at 70 DAF, i.e. at about 8 d after mass maturity. Thereafter, germination of fresh seeds declined, due to the imposition of a hard seed coat. Tolerance of desiccation to 10% moisture content was first detected at 56 DAF and was complete within the seed population by 84 DAF, i.e. about 22 d after mass maturity. Hardseededness began to be induced when seeds were dried to about 15% moisture content and below, with a negative logarithmic relation between hardseededness and moisture content below this value.

Desiccation effects on germination and vigor of King palm seeds

Determinou-se a tolerância ao dessecamento de sementes de Archontophoenix alexandrae (Wendl. & Drude) e identificou-se o teste de vigor mais sensível para avaliar a deterioração de sementes nessa espécie. Frutos maduros foram colhidos na coleção de palmeiras do Instituto Agronômico em Campinas. Os frutos despolpados foram transportados em embalagem impermeável para a UNESP em Botucatu, onde as sementes foram secas a diferentes intervalos de tempo. Foram avaliados teor de umidade, germinação, comprimento da plântula e condutividade elétrica. Os resultados sugerem que sementes de A. alexandrae são recalcitrantes, com alta porcentagem de germinação (acima de 67%) quando não desidratadas (47% de umidade). Teores de água inferiores a 31,5% reduziram significativamente a taxa de germinação (<52,5%). Perda total da capacidade germinativa foi observada em sementes com 15,1% de umidade. Dentre os testes de vigor empregados, aquele que avalia a condutividade elétrica da água de embebição foi o mais sensível para a identificação da deterioração da semente em resposta à dessecação progressiva do lote em estudo.

Plant spacing and pollen quantity on yield and quality of squash seeds

O rendimento e a qualidade de sementes de abóbora podem ser aprimorados por uma densidade populacional de plantas adequada, além da quantidade de pólen, que influi na qualidade da polinização e fertilização. Para este estudo, aplicaram-se nove tratamentos, que resultaram da combinação fatorial de três espaçamentos entre plantas (0,8 x 0,3; 0,8 x 0,6 e 0,8 x 0,9 m) com duas quantidades de pólen (50% de uma antera e uma antera inteira) e a polinização natural por insetos. Foram avaliados parâmetros de produção de frutos e sementes, além da qualidade física e fisiológica das sementes. O delineamento experimental adotado foi em blocos ao acaso com cinco repetições e dez plantas úteis por amostragem. O aumento do espaçamento entre plantas elevou o número médio de frutos maduros e a produção de sementes por planta. A produtividade de sementes foi diretamente proporcional à quantidade de pólen usada durante a polinização. Já o fator espaçamento entre plantas não afetou a produção de sementes por área. A maior quantidade de pólen resultou em uma maior produção de sementes por planta e por área. A polinização manual, com a utilização de uma antera inteira, não diferiu da polinização natural com insetos, tanto para rendimento como para a qualidade de sementes.