CnMADS1, a MADS transcription factor, positively modulates cell proliferation and lipid metabolism in the endosperm of coconut (Cocos nucifera L.).

MAIN CONCLUSION: The function of the first MADS-box transcription factor from endosperm of coconut, CnMADS1, was characterized via seed-specific overexpression in Arabidopsis seeds and further confirmed in protoplasts of coconut. Coconut (Cocos nucifera L.), which belongs to the palm family (Arecaceae), is one of the world's most useful economical tropical crops. However, few genes related to coconut endosperm development have been studied. In previous research, an AGAMOUS-like (AGL) MADS-box transcription factor, named CnMADS1, was identified in the endosperm of coconut through the SSH cDNA library. In this paper, functional characterization of the CnMADS1 gene was carried out by seed-specific overexpression in A. thaliana seeds and protoplasts of coconut. The results indicated that in the twelve independent T2 transgenic Arabidopsis lines with high overexpression of CnMADS1, the size of the mature seeds of transgenic plants was increased significantly (19.64% increase in the long axis and 8.6% increase in the short axis) compared to that of the wild-type seeds. Moreover, the total lipid content also increased significantly in mature seeds of transgenic plants. After comparing the expression of related genes in wild-type and transgenic plants and confirmation by EMSA, AtOSR1, a regulatory gene related to seed size, was proven to be significantly up-regulated by CnMADS1 in transgenic plants. Moreover, the transient transformation of protoplasts of coconut also proved that CnLECRK3 (the homologous gene of AtOSR1 in coconut) is up-regulated by the CnMADS1 gene in the same way. All these results indicated that a similar regulation mode existed in Arabidopsis and the endosperm of coconut and ultimately affected the yield and quality of coconut copra.

Hierarchical Structure of the Cocos nucifera (Coconut) Endocarp: Functional Morphology and its Influence on Fracture Toughness.

In recent years, the biomimetic potential of lignified or partially lignified fruit pericarps has moved into focus. For the transfer of functional principles into biomimetic applications, a profound understanding of the structural composition of the role models is important. The aim of this study was to qualitatively analyze and visualize the functional morphology of the coconut endocarp on several hierarchical levels, and to use these findings for a more precise evaluation of the toughening mechanisms in the endocarp. Eight hierarchical levels of the ripe coconut fruit were identified using different imaging techniques, including light and scanning electron microscopy as well as micro-computer-tomography. These range from the organ level of the fruit (H0) to the molecular composition (H7) of the endocarp components. A special focus was laid on the hierarchical levels of the endocarp (H3-H6). This investigation confirmed that all hierarchical levels influence the crack development in different ways and thus contribute to the pronounced fracture toughness of the coconut endocarp. By providing relevant morphological parameters at each hierarchical level with the associated toughening mechanisms, this lays the basis for transferring those properties into biomimetic technical applications.

Proximate composition, extraction, characterization and comparative assessment of coconut (Cocos nucifera) and melon (Colocynthis citrullus) seeds and seed oils.

Proximate composition, extraction, characterization and comparative assessment of Cocos nucifera and Colocynthis citrullus seeds and seed oils were evaluated in this work using standard analytical techniques. The results showed the percentage (%) moisture, crude fibre, ash, crude protein, lipids and total carbohydrate contents of the seeds as 7.51 and 4.27, 7.70 and 5.51, 1.02 and 2.94, 10.57 and 11.67, 47.80 and 50.42 and 32.84 and 29.47 while the calorific values were 553.99 and 567.32 Kcal/100 g for C. nucifera and C. citrullus, respectively. The two seed oils were odourless and at room temperature (30 degrees C) liquids, with a pale yellow to yellowish colouration. Lipid indices of the seed oils indicated the Acid Values (AV) as 2.06-6.36 mg NaOH g(-1) and 2.99-6.17 mg NaOH g(-1), Free Fatty Acids (FFA) as 1.03-3.18 and 1.49-3.09%, Saponification Values (SV) as 252.44-257.59 and 196.82-201.03 mg KOH g(-1), Iodine Values (IV) as 9.73-10.99 and 110.93-111.46 mg of I2 g(-1) of oil and Peroxide Values (PV) as 0.21-0.21 and 1.53-2.72 mg O2 kg(-1) for soxhlet-mechanical extracted C. nucifera and C. citrullus seed oils, respectively. The studied characteristics of the oil extracts in most cases compared favourably with most conventional vegetable oils sold in the Nigeria markets; however, there were some observed levels of significant differences in the values at p < or = 0.05. These results suggest that the seeds examined may be nutritionally potent and also viable sources of seed oils judging by their oil yield. The data also showed that the seed oils were edible inferring from their low AV and their corresponding low FFA contents. Industrially, the results revealed the seed oils to have great potentials in soap manufacturing industries because of their high SV. They were also shown to be non-drying due to their low IV which also suggested that the oils contain few unsaturated bonds and therefore have low susceptibility to oxidative rancidity and deterioration as confirmed by their low PV which also serves as indicators of the presence or high levels of anti-oxidants in the oils.

Cryopreservation of coconut (Cocos nucifera L.) zygotic embryos does not induce morphological, cytological or molecular changes in recovered seedlings.

The present study aimed at exploring the fidelity of coconut (Cocos nucifera L.) plants recovered from cryopreservation. Zygotic embryos from various different cultivars were cryopreserved following four successive steps, namely: rapid dehydration, rapid freezing, rapid thawing and in vitro recovery followed by acclimatization. At the end of the acclimatization period, the seedlings were compared to counterparts of the same age, which were produced from non-cryopreserved embryos. Both series were submitted to morphological, cytological and molecular comparisons. No significant differences in terms of growth rates could be measured. In addition, no morphological variation could be detected through the measurement of shoot elongation rates, production of opened leaves, and the number and total length of primary roots. Karyotype analysis revealed the same chromosome number (2n = 32) in all studied cultivars independently of cryopreservation. No significant differences could be observed between control and cryopreserved material concerning the type of chromosomes, the length of the long and short arms, the arm length ratio and the centromeric index. However, idiogram analysis did show a greater number of black banding on chromosomes isolated from cryopreserved material. Genetic and epigenetic fidelity was assessed through microsatellite (SSR) analysis and global DNA methylation rates; no significant differences would be observed between genomic DNAs isolated from seedlings originating from cryopreserved embryos and respective controls. In conclusion, our results suggest that the method of cryopreservation under study did not induce gross morphological, genetic or epigenetic changes, thus suggesting that it is an appropriate method to efficiently preserve coconut germplasm.

Accumulation of phenylpropanoid derivatives in chitosan-induced cell suspension culture of Cocos nucifera.

Chitosan-induced elicitation responses of dark-incubated Cocos nucifera (coconut) endosperm cell suspension cultures led to the rapid formation of phenylpropanoid derivatives, which essentially mimics the defense-induced biochemical changes in coconut palm as observed under in vivo conditions. An enhanced accumulation of p-hydroxybenzoic acid as the major wall-bound phenolics was evident. This was followed by p-coumaric acid and ferulic acid. Along with enhanced peroxidases activities in elicited lines, the increase in activities of the early phenylpropanoid pathway enzymes such as, phenylalanine ammonia lyase (PAL), p-coumaroyl-CoA ligase (4CL) and p-hydroxybenzaldehyde dehydrogenase (HBD) in elicited cell cultures were also observed. Furthermore, supplementation of specific inhibitors of PAL, C4H and 4CL in elicited cell cultures led to suppressed accumulation of p-hydroxybenzoic acid, which opens up interesting questions regarding the probable route of the biosynthesis of this phenolic acid in C. nucifera.

Correlation between the distribution of lignin and pectin and distribution of sorbed metal ions (lead and zinc) on coir (Cocos nucifera L.).

Plant fibres are capacious for sorption of metal ions, and can be used in water cleaning. Knowledge about the sorption will help in selection of the fibre and optimisation of its chemical modification, if any. The aim of this paper is to investigate the connection, if any, between the distribution of lignin and pectin and the loading of Pb and Zn on coir (mesocarp fibres from Cocos nucifera L.). The coir consisted mainly of xylem and a fibre sheath. The lignin was evenly distributed in the cell walls of the fibre sheath, but in the xylem, there was no detectable content in the compound middle lamella, and a smaller content of lignin in the secondary walls than in the walls of the fibre sheath. The only detectable content of pectin in the fibre sheath walls was in the middle lamella, cell corners and extracellular matrix, while in the xylem, the pectin was almost evenly distributed in the wall, with a higher concentration in the middle lamella and cell corners. All cell walls facing the lacuna had a high content of pectin. The metal ions were mainly loaded on the xylem and cell walls facing the lacuna, maybe with an additional trend to be loaded on the large fibres. Lead was distributed on and across the whole secondary wall. Zinc was loaded on the secondary walls, but there was no information about the distribution across the wall. If there is a simple correlation between the loading of metal ions and the distribution of lignin or pectin, these investigations point at no correlation with lignin and a positive correlation with pectin. It has to be stressed that these conclusions are made on limited material and are therefore preliminary in nature.

Cryopreservation by encapsulation-dehydration of plumules of coconut (Cocos nucifera L.).

This study describes the use of an encapsulation-dehydration cryopreservation technique on coconut plumules (apical dome with three or four leaf primordia) excised from embryos. In order to establish a reliable cryopreservation process for plumules, several different key factors were tested: pretreatment duration, sugar concentration, dehydration period and freezing. In parallel, histological studies were performed to describe the structural changes of tissues and plumule cells subjected to dehydration and freezing. A good survival level of around 60% was obtained. However, after 8 months culture regrowth, this level decreased to a maximum of 20 % which was achieved using sucrose treatment. In this paper we report for the first time the regeneration of leafy shoots from coconut plumules after cryopreservation.

Unfertilized ovary: a novel explant for coconut (Cocos nucifera L.) somatic embryogenesis.

Unfertilized ovaries isolated from immature female flowers of coconut (Cocos nucifera L.) were tested as a source of explants for callogenesis and somatic embryogenesis. The correct developmental stage of ovary explants and suitable in vitro culture conditions for consistent callus production were identified. The concentration of 2,4-dichlorophenoxyacetic acid (2,4-D) and activated charcoal was found to be critical for callogenesis. When cultured in a medium containing 100 microM 2,4-D and 0.1% activated charcoal, ovary explants gave rise to 41% callusing. Embryogenic calli were sub-cultured into somatic embryogenesis induction medium containing 5 microM abscisic acid, followed by plant regeneration medium (with 5 microM 6-benzylaminopurine). Many of the somatic embryos formed were complete with shoot and root poles and upon germination they gave rise to normal shoots. However, some abnormal developments were also observed. Flow cytometric analysis revealed that all the calli tested were diploid. Through histological studies, it was possible to study the sequence of the events that take place during somatic embryogenesis including orientation, polarization and elongation of the embryos.

New biotechnological applications of coconuts

Coconut is a major crop of many poor nations. The present paper shows with inexpensive fluorescence microscopy of the enhanced green fluorescent protein (EGFP), that coconut water has the capability of synthesizing proteins from recombinant DNA vectors. After only 4 hrs of 100 pM application of the EGFP plasmid (pEGFP), a strong fluorescent signal was detected with an off-the-shelf, low-tech CCD webcam. Since natural coconut water is sterile, this heat-stable delivery system may be a good option for the dissemination of food supplements and pharmaceuticals in poor nations. As expensive substances could be expressed in coconut water, through bioreactor technology or micropropagation, the concepts presented here may prove invaluable to the economic and social advancement of many developing countries.