Oil Palm (Elaeis guineensis Jacq.) Clonal Fidelity: Endogenous Cytokinins and Indoleacetic Acid in Embryogenic CallusCultures

Endogenous levels of indole-3-acetic acid (I AA) and cytokininswere measured in the two types of callus of oil palm (ElaeisguineensisJacq.) by an HPLC-ELISA methodology. The Nodular Compact Callus(NCC) is commonly used to establish stableembryoid strains ensuringclonal fidelity(     

In Vitro Culture of Rudimentary Embryos of Ilex paraguariensis: Responses to Exogenous Cytokinins

The effects of benzyladenine (BAP), kinetin (KIN), zeatin (ZEA), isopentenyladenine (2iP), and thidiazuron (TDZ) were studied on in vitro growth of rudimentary embryos of Ilex paraguariensis St. Hil. Heart stage zygotic embryos were removed from seeds of immature, light green fruits and cultured aseptically on quarter-strength Murashige and Skoog medium containing 3% sucrose, 0.65% agar, and supplemented with or without three concentrations of BAP, KIN, ZEA, 2iP, or TDZ. Cultures were incubated in darkness at 27 ± 2°C. Media containing 4.4 × 10⁻⁶ m BAP, 4.6 × 10⁻⁶ m KIN, or 4.9 × 10⁻⁶ m 2iP were totally ineffective in inducing embryo growth after culture for 28 days. However, lower concentrations of these compounds (4.4 × 10⁻⁸ m BAP, 4.6 × 10⁻⁸ m KIN, 4.5 × 10⁻⁸ m ZEA, or 4.9 × 10⁻⁸ m 2iP) promoted embryo growth. TDZ at 9.9 × 10⁻⁹ m, 9.9 × 10⁻⁸ m, or 9.9 × 10⁻⁷ m induced embryo growth at similar rates. The maximum percentage of embryos converted to seedlings was achieved when the medium was supplemented with 4.5 × 10⁻⁷ m ZEA. Received August 1, 1997; accepted February 19, 1998     

Variation in oil palm (Elaeis guineensis Jacq.) tissue culture-derived regenerants revealed by AFLPs with methylation-sensitive enzymes

Tissue culture-derived plants of oil palm (Elaeis guineensis Jacq.) can develop abnormal flowers in which stamen primordia are converted into carpel-like tissues (mantled fruit). This abnormality can be heritable; individual palms may show variation in mantling and reversion to the normal phenotype over time has been observed. Four sets of ortets (mother plant used as tissue source) and ramets (regenerated plants) were compared using standard amplified fragment length polymorphism (AFLP) analysis and AFLPs using methylation-sensitive enzymes. No polymorphisms were found when standard AFLPs were produced with ten different primer combinations. In contrast, when methylation-sensitive AFLPs were used, polymorphisms were detectable. Polymorphisms appeared as new bands in the ramets, suggesting that a reduction in methylation had occurred during tissue culture. The highest number of polymorphic bands (0.3%) was obtained when HpaII was used as the restriction endonuclease, indicating that the loss of methylation had occurred most frequently at the internal C within the HpaII recognition sequence 5’-CCGG-3’. Conversion of nine of the polymorphic bands into probes for Southern analysis confirmed that these were not due to partial digestion of the AFLP templates and showed that the majority were single-copy sequences. The exceptions were fragments showing homology to 25S ribosomal RNA genes and the chalcone synthase gene family. Examination of the Southern blots suggested that most of the single-copy sequences were partially de-methylated, and one example was found in which de-methylation affected only one allele. No polymorphism was consistently different between normal and abnormal clones in all the sets. This suggests that, whilst this method is an effective way of detecting variation in tissue culture-derived plants, different approaches will be required to identify the causal basis of the mantled fruit abnormality. Received: 25 May 2000 / Accepted: 28 August 2000     

Cytokinin Differences in In Vitro Cultures and Inflorescences from Normal and Mantled Oil Palm (Elaeis guineensis Jacq.)

The mantled abnormality phenotype of the oil palm affects fruit development and thus jeopardizes oil yield. Cytokinins have been implicated in the development of the mantled phenotype. Endogenous cytokinin levels in the normal and mantled phenotypes were compared to determine whether levels of specific cytokinins are associated with mantling. Endogenous cytokinins were identified and quantified in in vitro cultures and inflorescences from normal and mantled oil palms. Twenty-two isoprenoid cytokinins, comprising the zeatin, dihydrozeatin, and isopentenyladenine types, were quantified. Total cytokinin levels, particularly of trans-zeatin and isopentenyladenine types, increased during the in vitro culture process, with the highest levels detected at the proliferating polyembryoid stages. The cytokinins were present mainly in their inactive 9-glucoside forms during in vitro culture. On the other hand, the predominant trans-zeatin cytokinins in inflorescences were present mainly in their ribotide forms, suggesting a metabolic pool of cytokinins for conversion to biologically active free bases or ribosides. Levels of specific cytokinins were significantly different in tissues at different stages. Mantled developed inflorescences contained higher levels of isopentenyladenine 9-glucoside compared with normal inflorescences. Mantled-derived callus tissues had higher isopentenyladenine levels but significantly lower levels of trans-zeatin 9-glucoside, dihydrozeatin riboside, and dihydrozeatin riboside 5′-monophosphate cytokinins compared with normal-derived callus. It would be of considerable interest to verify these specific cytokinin differences in more callus cultures and clones.     

High-temperature Pre-treatment and the Germination of Seed of the Oil Palm, Elaeis guineensis (Jacq.)

Germination experiments have shown that oil palm seeds requireheattreatment before they germinate. The heat-treatment is insome ways analogous to low-temperature stratification of temperateseed, as it may be applied as a pretreatment, even at a moisturecontent too low for germination to occur. Following the heatpre-treatment, seed brought to the optimum moisture contentgerminates rapidly, after an initial lag period, at ambienttemperature. Below a critical seed moisture content, heat-treatmentis ineffective although seed viability is largely unimpaired.The high-temperature reaction has a Q₁₀ between 3.5 and 5.0and is apparently irreversible. Heat-treatment may be effectivelyapplied to stored, as well as fresh, seed. A reinvestigationof the optimum temperature for the high-temperature reactionshowed that dry heat-treatment is effective at 42.0°C.,the duration of the treatment necessary being only 60 days comparedwith the previously accepted value of 80 days at 39.5°C.necessary for seed heat-treated at optimum moisture content.A temperature of 44.5°C. is rapidly fatal. These resultsare discussed in relation to the alternation of dry and wetseasons in West Africa, and in relation to practical aspectsof oil palm cultivation.     

Composition, Degradation and Utilization of Endosperm During Germination in the Oil Palm (Elaeis guineensis Jacq.)

The insoluble carbohydrate and lipid fractions, and -D-galactosidase,ß-D-mannosidase and isocitrate lyase activities werestudied in the various tissues of oil palm (Elaeis guineensisJacq.) kernels prior to and during germination. In ungerminatedkernels insoluble carbohydrate and lipid constituted 36 and47% of endosperm dry weight respectively. During germinationthe thick endosperm cell walls became markedly thinner, concurrentwith a significant decrease in the percentage of insoluble carbohydrateand an increase in -galactosidase and ß-mannosidaseactivity in both degraded and residual endosperm. The proportionof lipid in degraded endosperm also increased significantly.The insoluble carbohydrate appears to be a galactomannan locatedin the secondary walls of the endosperm. No galactomannan wasdetected in oil palm embryos or haustoria. Isocitrate lyasewas present in, and confined to, tissues of the haustorium ofgerminating kernels. The enzyme was not active in endospermat any stage of germination, nor was it active in embryos beforeor at the end of imbibition. The results suggest that galactomannan is the second largestcomponent of oil palm endosperm and that it is utilized morerapidly than lipid during the early stages of germination. Thefact that isocitrate lyase activity is confined to the haustoriumsuggests that in Elaeis gluconeogenesis, the conversion of triglycerideto carbohydrate, takes place entirely within the cotyledon ofthe seed. Elaeis guineensis, galactomannan, galactosidase, germination, isocitrate lyase, mannosidase, oil palm     

An Analysis of the Factors Controlling the Germination of the Seed of the Oil Palm, Elaeis guineensis (Jacq.)

Under natural conditions in West Africa at the average groundtemperatures of 25-30° C, oil-palm seed germinates sporadicallyover a period of years, but with continuous heat treatment (38-40°C.) 50 per cent, germination of either nuts or extracted kernelsoccurs within a few months. The kernel comprises a small cylindrical embryo embedded ina mass of oily endosperm which in turn is surrounded by a thicktesta. At germination the testa and layer of endosperm coveringthe distal part of the embryo are forced out as a disc-likeoperculum which is already demarcated in the ungerminated kernelby a circular abscission zone in the endosperm cells. The excised embryo is non-dormant and at 30° C. starts toelongate within 24 hours, either on moist filter paper or whilestill in contact with the endosperm. At 30° C. removal of the operculum is followed by sluggishemergence of the embryo in air; emergence at the normal rateoccurs in pure oxygen, or in air if a long period at high temperatureprecedes de-operculation. Fresh intact kernels remain ungerminatedfor at least 6 months at 30° C, either in air or in pureoxygen. At high temperature (40° C.) intact kernels germinate fasterin pure oxygen than in air; at 30° C. the same applies onlyafter previous treatment at high temperature. This acceleratingeffect of oxygen is shown to be dependent upon the progressof slow processes stimulated by high temperature. It is postulated that germination of oil-palm seed is dependentupon a minimal threshold concentration of oxygen in the embryoand that during dormancy this minimal level progressively decreasesat a rate depending on the temperature.     

Assessment of nuclear,mitochondrial and chloroplast RFLP markers in oil palm (Elaeis guineensis Jacq.)

A variety of DNA probes was used to screen a diverse set of oil palm accessions in order to identify markers with a utility in genotype discrimination. This survey included samples of the commercial oil palm native to Africa (Elaeis guineensis Jacq.), the closely-related South American species [E.oleifera (HBK) Cortes] and inter-specific hybrids of the two. Of 106 major chloroplast bands none showed differences between E. guineensis and E. Oleifera. Mitochondrial and ribosomal probes were more informative inter-specifically (the former allowing identification of the maternal inheritance of mitochondria) and may be useful in hybrid breeding programmes; however, they were unable to identify polymorphism within E. guineensis. In contrast, low-copy nuclear genomic clones were able to identify intra-specific variation, though in most cases they revealed a relatively small number of allelic variants. One DNA probe showed a much larger number of band variants, revealing ten patterns amongst 13 E. guineensis accessions, and should prove useful in genetic fingerprinting and evaluation of oil-palm germplasm collections.     

Expression Analysis of Fatty Acid Biosynthetic Pathway Genes during Interactions of Oil Palm (Elaeis guineensis Jacq.) with the Pathogenic Ganoderma boninense and Symbiotic Trichoderma harzianum Fungal Organisms

The fatty acid (FA) signaling pathway is emerging as an important mechanism in plant responses during interactions with microbial organisms. For a comprehensive evaluation of key FA biosynthetic pathway genes during interactions of oil palm (Elaeis guineensis Jacq.) with the pathogenic Ganoderma boninense and symbiotic Trichoderma harzianum fungal organisms, a lane-based array analysis of gene expression in artificially inoculated oil palm seedlings was performed. The results obtained demonstrated that acetyl-CoA carboxylase (ACC), β-ketoacyl-ACP synthases (KAS) II and III, palmitoyl-ACP thioesterase (PTE), oleoyl-ACP thioesterase (OTE) and glycerol-3-phosphate acyltransferase (ACT) showed identical responses in root and leaf tissues for the same fungi. The expression of these genes was up-regulated in both root and leaf tissues at 21 days post-inoculation (dpi) during interaction of oil palm with G. boninense. Thereafter, production of physical symptoms occurred at 42 and 63 dpi concomitantly with suppression of expression of these genes. An increase in the expression level of these genes was observed in both tissues at 3–63 dpi, which correlated with the colonization of roots and promotion of plant growth by T. harzianum. These data suggest that FA biosynthetic pathway genes are involved in the defense response of oil palm to infection. Identical plant responses by FA biosynthetic pathway genes may lead to enhanced resistance against G. boninense and could be a useful marker to contribute towards early detection of infection. The distinct expression profile during symbiotic interaction demonstrated its role in plant resistance mechanisms and growth promotion by T. harzianum.     

Effects of Chemical Growth Retardants on Growth and Development of Sweetpotato (Ipomoea batatas (L.) Lam.) in Vitro

Plant growth retardants were evaluated for their ability to reduce the growth rate of sweetpotato (Ipomoea batatas (L.) Lam.) in vitro. Nodal sections of cv. Jewel were cultured for 30 days on medium containing NDA, ancymidol, phosfon, TIBA, difenzoquat, chlormequat, ACC, mepiquat chloride, or daminozide at 0, 10⁻⁴, 10⁻⁵, 10⁻⁶, 10⁻⁷, or 10⁻⁸ m. Difenzoquat, NDA, phosfon, and TIBA, at 10⁻⁴ m, were lethal to axillary bud explants. A low concentration (10⁻⁸ m) of chlorflurenol or NDA stimulated shoot elongation. The effective concentration range for most growth retardants was 10⁻⁵ to 10⁻⁶ m. Small (2- to 4-mm diameter) storage root-like swellings were observed on roots in cultures containing TIBA or ancymidol. The growth-inhibiting effects of ancymidol and NDA were transitory and did not persist through a 180-day culture period. Shoots cultured on medium containing 10⁻⁵ m phosfon, TIBA, or difenzoquat were significantly shorter than control plants after a 180-day culture period. Culture on medium containing TIBA, NDA, ancymidol, or ACC resulted in abnormal leaf and stem development. Plants derived from nodal explants cultured on medium containing either phosfon or chlormequat were near normal in appearance but with some plants exhibiting interveinal chlorosis and reduced root system development. Received May 9, 1997; accepted August 14, 1997     

A new aspect of genetic diversity of Indonesian oil palm (Elaeis guineensis Jacq.) revealed by isoenzyme and AFLP markers and its consequences for breeding

Oil palm (Elaeis guineensis Jacq.) plays an important economic role in some countries of Southeast Asia like Indonesia, which is the world’s second producer of palm and palm kernel oil. The quality improvement of planting material needs a better understanding of the genetic relationships between genotypes from different populations used in the breeding programmes. In this study, 48 parents, representative of four populations used in Indonesian Oil Palm Research Institute (IOPRI) breeding programmes, were analysed with five selected AFLP primer pairs and four isoenzymatic systems. One hundred and fifty eight scorable band levels were generated of which 96 (61%) were polymorphic. AFLP allowed us to identify off–type descendants which were excluded from analysis. The use of unbiased Rogers distance clearly separated the four studied populations. The Neighbor-Joining method re-groups two African populations which are known as originating from different regions. Nevertheless, the variability revealed is in accordance with oil palm breeders’ knowledge. The results obtained with AFLP showed that the crosses among the African sub-population, which is excluded in oil palm reciprocal recurrent selection (RRS) breeding programmes, may be more interesting than the crosses between the African and the Deli populations. Received: 9 August 1999 / Accepted: 28 February 2000     

Sequence Analysis of 16S rRNA and secA Genes Confirms the Association of 16SrI‐B Subgroup Phytoplasma with Oil Palm (Elaeis guineensis Jacq.) Stunting Disease in India

During January 2010, severe stunting symptoms were observed in clonally propagated oil palm (Elaeis guineensis Jacq.) in West Godavari district, Andhra Pradesh, India. Leaf samples of symptomatic oil palms were collected, and the presence of phytoplasma was confirmed by nested polymerase chain reaction (PCR) using universal phytoplasma‐specific primer pairs P1/P7 followed by R16F2n/R16R2 for amplification of the 16S rRNA gene and semi‐nested PCR using universal phytoplasma‐specific primer pairs SecAfor1/SecArev3 followed by SecAfor2/SecArev3 for amplification of a part of the secA gene. Sequencing and BLAST analysis of the ～1.25 kb and ～480 bp of 16S rDNA and secA gene fragments indicated that the phytoplasma associated with oil palm stunting (OPS) disease was identical to 16SrI aster yellows group phytoplasma. Further characterization of the phytoplasma by in silico restriction enzyme digestion of 16S rDNA and virtual gel plotting of sequenced 16S rDNA of ～1.25 kb using iPhyClassifier online tool indicated that OPS phytoplasma is a member of 16SrI‐B subgroup and is a ‘Candidatus Phytoplasma asteris’‐related strain. Phylogenetic analysis of 16S rDNA and secA of OPS phytoplasma also grouped it with 16SrI‐B. This is the first report of association of phytoplasma of the 16SrI‐B subgroup phytoplasma with oil palm in the world.     

Conditioning of Fasciation by Gibberellin and Genotype in Cotton (Gossypium hirsutum L.)

The round-leafed mutant cotton line L-501 developed fasciation of the upper stem when field grown in Central Asia. Fasciation co-segregated with the mutant gene for round leaves In. ^l Fasciation developed at the flowering stage, but removal of floral buds did not prevent fasciation. Fasciation in L-501 could be inhibited by the gibberellin (GA) biosynthesis inhibitor chlorocholine chloride or by fusicoccin. GA₃ application in the field induced fasciation in the mutant's parental line L-463, which has five-lobed leaves and does not normally develop fasciation. Fasciation did not develop in either line, even after GA₃ treatment, in UK glasshouse conditions. Received June 17, 1998; accepted January 25, 1999     

Effects of Waterlogging on Nitrogen Accumulation and Alleviation of Waterlogging Damage by Application of Nitrogen Fertilizer and Mixtalol in Winter Rape (Brassica napus L.)

A study on the physiological and yield effects of waterlogging and the alleviation of waterlogging damage by the application of nitrogen fertilizers and mixtalol in winter rape was conducted in experimental tanks especially designed for controlling soil moisture content. The results showed that waterlogging at the seedling and stem elongation stages causes a significant decrease in nitrogen content and rate of nitrogen accumulation. Leaf chlorophyll content, superoxide dismutase and catalase activities, and root oxidizability (capacity for root oxidation) and root exudate were also reduced by waterlogging. The experiments confirmed that the physiological function of rape plants was retarded during the time of waterlogging at the seedling stage, and its adverse effects remained. Plant height, stem width, and the number of primary branches per plant were decreased significantly by waterlogging at the seedling and stem elongation stages. Pods per plant and seeds per pod were also reduced significantly, giving a 21.3 and 12.5% decrease of seed yield from the control for treatments at the seedling and stem elongation stages, respectively. Foliar sprays of nitrogen fertilizers at the seedling stage or mixtalol at the flowering stage alleviated plant damage caused by waterlogging by retarding chlorophyll and nitrogen degradation, increasing superoxide dismutase and catalase activities and root oxidizability, and improving yield components and seed yield of waterlogged plants. Therefore, besides draining off water, alleviation of waterlogging damage may be controlled by applying nitrogen fertilizer and a suitable plant growth regulator at appropriate growth stages. Received July 3, 1996; accepted December 26, 1996     

红菜薹游离小孢子培养与植株再生

以5个红菜薹(Brassica compestris ssp．chinensis var．pupurea Hort．)基因型为试材,探讨了基因型和活性炭对产胚量的影响。结果表明：产胚量最高的是基因型8902,达到42个／皿,最少的为零;加适量活性炭可以使产胚量提高近3倍。同时,对胚状体进一步再生成苗因素也进行了研究：在培养基中添加1．2％的琼脂浓度再生率最高,达到50．1％;4℃下处理10d可使再生成苗率从45％提高到65％;随胚状体年龄的延长,其再生成株率明显降低,最适的胚龄是20-24d;而培养基B5和MS对小孢子再生率的影响不大。     

Association of Copper and Zinc Levels in Oil Palm (Elaeis guineensis) to the Spatial Distribution of Ganoderma Species in the Plantations on Peat

Nutrients are essential for normal physiological processes in plants, and they play important roles in defence mechanisms against pathogens. Oil palms cultivated on peat are more prone to nutrient deficiency, especially micronutrients, and this may affect their susceptibility to Ganoderma species, the major threat to the sustainability of oil palm throughout South‐East Asia. This study was conducted to investigate the association of copper (Cu) and zinc (Zn) in mature oil palm to the spatial distribution of Ganoderma species in the plantations on peat. Foliar samples (frond 17) of oil palm from two plantations (Betong and Miri) on peat in Sarawak, Malaysia, were collected based on the spatial distribution pattern of Ganoderma, and total Cu and Zn were quantified spectrometrically. The experiment was conducted twice at a 1‐year interval. The concentrations of Cu and Zn were significantly lower in oil palms from infected areas in contrast to those from uninfected areas. In addition, oil palms in infected areas in Miri suffered Cu and Zn deficiencies. Furthermore, Cu and Zn were significantly lower in the oil palms in Miri that had higher Ganoderma occurrence, as compared to those in Betong, which had significantly higher Cu and Zn but lower Ganoderma occurrence.