Feasibility of enzymatic hydrolysis and alcoholic fermentation of starch contained in buffalo gourd (Cucurbita foetidissima) roots

The suitability of using annually grown, carrot-sized buffalo gourd (Cucurbita foetidissima) roots as a feedstock for alcoholic fermentation was explored. Roots grown in 1982 and 1983 were slurried, dextrinized and saccharified using Takathermtrade mark and Diazymetrade mark (commercial enzymes manufactured by Miles Laboratories), and fermented by the action of Saccharomyces cerevisiae. These processes were monitored in detail and results were compared with those displayed by controls formulated using potato tubers. The preparation of gourd root slurries with suitable viscosity characteristics for enzymatic digestion required the addition of water (at least 50% by weight) which reduced the proportion of fermentable sugars in the resulting saccharified suspensions. The resulting slurries were well-suited to enzymatic conversion of starch to sugar. Estimates of enzymatic efficiency in gourd root suspensions did not suggest the presence of naturally occurring amylase or glucosidase inhibitors in these plant materials. Saccharified gourd root mashes supported yeast growth well and produced ethanol yields at 82.2-86.5% of the theoretically maximum efficiency.     

Pollen competition in a natural population of Cucurbita foetidissima (Cucurbitaceae)

The pollen competition hypothesis predicts that when the number of pollen grains deposited onto stigmas exceeds the number of ovules, selection can operate in the time frame between deposition and fertilization. Moreover, because of the overlap in gene expression between the two phases of the life cycle, selection on microgametophytes may alter the resulting sporophytic generation. The extent to which pollen competition occurs in nature has been unclear, because tests of the predictions of the pollen competition hypothesis have used cultivars and/or artificial growth conditions and hand-pollination techniques. In this study we used a wild species, Cucurbita foetidissima, in its natural habitat (southern New Mexico) to determine the amount and timing of the arrival of pollen onto stigmas, the relationship between pollen deposition and seed number, and the effects of the intensity of pollen competition on progeny vigor. We found that ～900 pollen grains are necessary for full seed set and that a single visit by a pollinator results in the deposition of 653.0 ± 101.8 pollen grains. About 29% of the flowers receiving a single pollinator visit had 900 or more pollen grains on its stigma. Moreover, within 2 h of anthesis, >4000 pollen grains were deposited onto a typical stigma, indicating that multiple pollinator visits must have occurred. Fruits produced by multiple visits had greater seed numbers (206 vs. 147) than fruits produced by a single visit. Finally, the progeny produced by multiple pollinator visits were more vigorous than those produced by single visits with respect to five measures of vegetative growth (MANCOVA, Wilks' lambda = 0.96, F(6,370) = 2.54, P < 0.02. These data demonstrate that conditions for pollen competition exist in nature and support the prediction that pollen competition enhances offspring vigor.     

Experimental investigation of pathogenic stress on phytolith formation in Cucurbita pepo var. texana (wild gourd)

Silica phytoliths that form in plant tissues are useful to archaeologists because of their diagnostic value and longevity in ancient deposits. Palaeoecology, site formation processes, plant domestication, and other topics are routinely addressed using phytolith assemblages, especially when macrobotanical remains are not well preserved. However, little research has been conducted to document the effects of ecological variables on phytolith formation. Here, we investigate the effects of mosaic virus and bacterial wilt disease on diagnostic scalloped phytoliths in the rind of a wild-type Cucurbita pepo var texana (gourd). We observe a minimal change in phytolith size distribution between control plants and individuals with mosaic virus. However, we observe a notable difference between plants with bacterial wilt disease and control plants, with diseased individuals carrying a greater proportion of large-diameter scalloped phytoliths. This and similar phenomena could potentially confound archaeological interpretations of phytolith assemblages, and we suggest that the effects of this and other ecological variables should be studied in a diverse range of taxa.     

Transferability of Cucurbita SSR markers for genetic diversity assessment of Turkish bottle gourd (Lagenaria siceraria) genetic resources

The genetic diversity present in crop landraces represents a valuable genetic resource for breeding and genetic studies. Bottle gourd (Lagenaria siceraria) landraces in Turkey are highly genetically diverse. However, the limited genomic resources available for this crop hinder the molecular characterization of Turkish bottle gourd germplasm for its adequate conservation and management. Therefore, we evaluated the efficacy of 40 SSR markers from major cucurbit crops (Cucurbita pepo L. and Cucurbita moschata L.) in 30 bottle gourd landraces, together with 16 SRAP primer combinations. In addition, we compared the genetic relationship between bottle gourd and 31 other cucurbit accessions (11 Cucurbita maxima, 3 C. moschata, 5 C. pepo subsp. ovifera, 10 C. pepo and 2 Luffa cylindrica). Twenty-seven Cucurbita SSR markers showed transferability to bottle gourd. SSR markers amplified 59 alleles, in bottle gourd genome with an average of 1.64 alleles per locus. Together, SSR and SRAP markers amplified 453 fragments across the 61 accessions, and clearly discriminated L. siceraria and L. cylindrica from the other cucurbit species. Genetic diversity analysis separated edible cucurbit from ornamentals, while population structure analysis classified L. siceraria in two subpopulations defined by fruit shape, rather than geographical origin. The results indicated that the genomic resources available for Cucurbita species are valuable to study and preserve the genetic diversity of bottle gourd in Turkey.     

Light-induced transpiration alters cell water relations in figleaf gourd (Cucurbita ficifolia) seedlings exposed to low root temperatures

Water relation parameters including elastic modulus (epsilon), half-times of water exchange (T(w)(1/2)), hydraulic conductivity and turgor pressure (P) were measured in individual root cortical and cotyledon midrib cells in intact figleaf gourd (Cucurbita ficifolia) seedlings, using a cell pressure probe. Transpiration rates (E) of cotyledons were also measured using a steady-state porometer. The seedlings were exposed to low ambient (approximately 10 micromol m(-2) s(-1)) or high supplemental irradiance (approximately 300 micromol m(-2) s(-1) PPF density) at low (8 degrees C) or warm (22 degrees C) root temperatures. When exposed to low irradiance, all the water relation parameters of cortical cells remained similar at both root temperatures. The exposure of cotyledons to supplemental light at warm root temperatures, however, resulted in a two- to three-fold increase in T(w)(1/2) values accompanied with the reduced hydraulic conductivity in both root cortical (Lp) and cotyledon midrib cells (Lp(c)). Low root temperature (LRT) further reduced Lp(c) and E, whether it was measured under low or high irradiance levels. The reductions of Lp as the result of respective light and LRT treatments were prevented by the application of 1 microM ABA. Midrib cells required higher concentrations of ABA (2 microM) in order to prevent the reduction in Lp(c). When the exposure of cotyledons to light was accompanied by LRT, however, ABA proved ineffective in reversing the inhibition of Lp. LRT combined with high irradiance triggered a drastic 10-fold reduction in water permeability of cortical and midrib cells and increased epsilon and T(w)(1/2) values. Measurement of E indicated that the increased water demand by the transpiring plants was fulfilled by an increase in the apoplastic pathway as principal water flow route. The importance of water transport regulation by transpiration affecting the hydraulic conductivity of the roots is discussed.     

Detoxification of hydrogen peroxide maintains the water transport activity in figleaf gourd (Cucurbita ficifolia) root system exposed to low temperature

The figleaf gourd ( Cucurbita ficifolia Bouché) root system has the ability to take up water and nutrients at low soil temperatures, and in the present paper, we attempt to reveal some of the molecular mechanisms behind this low-temperature tolerance. Exposure of figleaf gourd root system to low temperature induced accumulation of H₂O₂ along the plasma membrane but not in the cytoplasm. H⁺-ATPase (EC 3.6.1.35) activity of isolated root plasma membranes and root hydraulic conductivity ( Lp_r ) were largely insensitive to externally applied H₂O₂. However, using bromocresol purple, it was shown that the acidification of the medium surrounding the root was strongly inhibited with low temperature- and H₂O₂-treated roots. Addition of catalase (EC 1.11.1.6) to the root medium during low-temperature exposure led to a recovery of H⁺-efflux along the root surface and increased Lp_r , demonstrating the importance of an H₂O₂ detoxification system in the root cells. Additional evidence for an increased Lp_r was obtained by the Fenton reaction wherein a warming of the solution increased the activity of the detoxification system. All available evidence suggests that the ability of figleaf gourd root system to maintain a low level of H₂O₂ in the cytoplasm and to detoxify reactive oxygen species is related to the maintenance of water transport activity at low temperatures.     

Development of ultra-high erucic acid oil in the industrial oil crop Crambe abyssinica

Erucic acid (22 : 1) is a major feedstock for the oleochemical industry. In this study, a gene stacking strategy was employed to develop transgenic Crambe abyssinica lines with increased 22 : 1 levels. Through integration of the LdLPAAT, BnFAE1 and CaFAD2-RNAi genes into the crambe genome, confirmed by Southern blot and qRT-PCR, the average levels of 18 : 1, 18 : 2 and 18 : 3 were markedly decreased and that of 22 : 1 was increased from 60% in the wild type to 73% in the best transgenic line of T4 generation. In single seeds of the same line, the 22 : 1 level could reach 76.9%, an increase of 28.0% over the wild type. The trierucin amount was positively correlated to 22 : 1 in the transgenic lines. Unlike high erucic rapeseed, the wild-type crambe contains 22 : 1 in the seed phosphatidylcholine and in the sn-2 position of triacylglycerols (5% and 8%, respectively). The transgenic line with high 22 : 1 had decreased 22 : 1 level in phosphatidylcholine, and this was negatively correlated with the 22 : 1 level at the sn-2 position of TAG. The significances of this study include (i) achieving an unprecedented level of 22 : 1 in an oil crop; (ii) disclosing mechanisms in the channelling of a triacylglycerol-specific unusual fatty acid in oil seeds; (iii) indicating potential limiting factors involved in the erucic acid biosynthesis and paving the way for further increase of this acid and (iv) development of an added value genetically modified oil crop having no risk of gene flow into feed and food crops.     

Guaraná (Paullinia cupana) as a commercial crop in Brazilian Amazonia

Guaraná (Paullinia cupana) is a woody vine or sprawling shrub native to the central Amazon Basin. The seeds are commercially produced on some 6,000 ha in the state of Amazonas near Manaus. The principal article of commerce is an amber-colored, carbonated soft drink. It is also widely used as a high caffeine stimulant and in local medicines. The plant is monoecious and is damaged by a number of diseases, the most severe being anthracnose. Prospects are excellent for greatly expanded international markets.     

Films of starch and poly(butylene adipate co-terephthalate) added of soybean oil (SO) and Tween 80

Starch extruded in the presence of a plasticizer results in a material called thermoplastic starch (TPS). TPS mixed with poly(butylene adipate co-terephthalate) (PBAT), soybean oil (SO), and surfactant may result in films with improved mechanical properties due to greater hydrophobicity and compatibility among the polymers. This study characterized films produced from blends containing 65% TPS and 35% PBAT with SO added as compatibilizer. The Tween 80 was added to prevention of phase separation. The elongation and resistance were greater in the films with SO. The infrared spectra confirmed an increase in ester groups bonded to the PBAT and the presence of groups bonded to the starch ring, indicating TPS-SO and PBAT-SO interactions. The micrographs suggest that the films with SO were more homogenous. Thus, SO is considered to be a good compatibilizer for blends of TPS and PBAT.     

南瓜高效再生体系的建立

南瓜(Cucurbita moschata)再生率较低, 为建立高效的南瓜再生体系, 以南瓜子叶为外植体, 进行35组不同激素浓度的不定芽诱导研究。结果表明, 南瓜再生受培养基中激素浓度和配比的影响, 适宜浓度6-苄氨基腺嘌呤(6-BA)能有效促进不定芽形成; 单独使用脱落酸(ABA)诱导使南瓜子叶发黄, 但与6-BA组合使用可显著提高外植体的再生能力, 1.0 mg∙L ^-16-BA与0.5 mg∙L ^-1ABA组合南瓜芽再生率高达90.26%。将不定芽置于MS培养基中进行生根培养, 再生苗移栽易成活。从子叶接种到苗再生约需70天。     

南瓜高效再生体系的建立

南瓜(Cucurbita moschata)再生率较低, 为建立高效的南瓜再生体系, 以南瓜子叶为外植体, 进行35组不同激素浓度的不定芽诱导研究。结果表明, 南瓜再生受培养基中激素浓度和配比的影响, 适宜浓度6-苄氨基腺嘌呤(6-BA)能有效促进不定芽形成; 单独使用脱落酸(ABA)诱导使南瓜子叶发黄, 但与6-BA组合使用可显著提高外植体的再生能力, 1.0 mg?L ^-16-BA与0.5 mg?L ^-1ABA组合南瓜芽再生率高达90.26%。将不定芽置于MS培养基中进行生根培养, 再生苗移栽易成活。从子叶接种到苗再生约需70天。     

Purification and characterization of a novel peroxidase from bitter gourd (Momordica charantia)

Peroxidase from bitter gourd was purified by three step purification scheme; ammonium sulphate fractionation, gel filtration and affinity chromatography. The enzyme was purified 42 fold with the retention of 67% of the initial activity. The enzyme exhibited its maximum activity at pH 5.6 and 40 degrees C. The enzyme retained half of its activity even after 1 h incubation at 60 degrees C. Molecular weight of the purified glycosylated bitter gourd peroxidase determined by Sephacryl S-100 and SDS-PAGE was 43 kDa. The stokes radius, diffusion coefficient and sedimentation coefficient of the purified peroxidase were 27.3 A, 8.17 x 10(-7) cm(2)/sec and 3.74 S, respectively. K(m) for o-dianisidine and ABTS were 1.3 and 4.9 mM, respectively. The activity of the enzyme was inhibited by sulfide, azide and L-cysteine. The carbohydrate content and sulfydryl groups of the enzyme were 25% (w/w) mass of the protein and 16 mmoles/mole of the protein, respectively.     

Calcium alginate–starch hybrid support for both surface immobilization and entrapment of bitter gourd (Momordica charantia) peroxidase

Calcium alginate–starch hybrid gel was employed as an enzyme carrier both for surface immobilization and entrapment of bitter gourd peroxidase. Entrapped crosslinked concanavalin A–bitter gourd peroxidase retained 52% of the initial activity while surface immobilized and glutaraldehyde crosslinked enzyme showed 63% activity. A comparative stability of both forms of immobilized bitter gourd peroxidase was investigated against pH, temperature and chaotropic agent; like urea, heavy metals, water-miscible organic solvents, detergent and inhibitors. Entrapped peroxidase was significantly more stable as compared to surface immobilized form of enzyme. The pH and temperature-optima for both immobilized preparations were the same as for soluble bitter gourd peroxidase. Entrapped crosslinked concanavalin A–bitter gourd peroxidase showed 75% of the initial activity while the surface immobilized and crosslinked bitter gourd peroxidase retained 69% of the original activity after its seventh repeated use.     

Genetic relationships and evolution in <Emphasis Type="Italic">Cucurbita pepo</Emphasis> (pumpkin,squash, gourd) as revealed by simple sequence repeat polymorphisms

Genetic relationships among 104 accessions of Cucurbita pepo were assessed from polymorphisms in 134 SSR (microsatellite) and four SCAR loci, yielding a total of 418 alleles, distributed among all 20 linkage groups. Genetic distance values were calculated, a dendrogram constructed, and principal coordinate analyses conducted. The results showed 100 of the accessions as distributed among three clusters representing each of the recognized subspecies, pepo, texana, and fraterna. The remaining four accessions, all having very small, round, striped fruits, assumed central positions between the two cultivated subspecies, pepo and texana, suggesting that they are relicts of undescribed wild ancestors of the two domesticated subspecies. In both, subsp. texana and subsp. pepo, accessions belonging to the same cultivar-group (fruit shape) associated with one another. Within subsp. pepo, accessions grown for their seeds or that are generalists, used for both seed and fruit consumption, assumed central positions. Specialized accessions, grown exclusively for consumption of their young fruits, or their mature fruit flesh, or seed oil extraction, tended to assume outlying positions, and the different specializations radiated outward from the center in different directions. Accessions of the longest-fruited cultivar-group, Cocozelle, radiated bidirectionally, indicating independent selection events for long fruits in subsp. pepo probably driven by a common desire to consume the young fruits. Among the accessions tested, there was no evidence for crossing between subspecies after domestication.     

Development and validation of a simple, sensitive enzyme immunoassay (EIA) for quantification of prolactin in buffalo plasma

A simple and highly sensitive enzyme immunoassay (EIA) was developed and validated for prolactin quantification in buffalo plasma (on a microtitreplate) using the biotin-streptavidin-peroxidase amplification and immobilized antiserum in a competitive assay. Prolactin standards (range: 5-5000 pg/(well 50 microL)) were prepared in hormone-free plasma collected from minimal stress non-lactating buffalo heifers in temperate weather. The sensitivity of the EIA procedure was 5 pg/(well 50 microL) (corresponds to 0.1 ng/mL plasma); the 50% relative binding sensitivity occurred at 160 ng/(well 50 microL). Plasma volumes for the EIA, viz. 12.5, 25, and 50 microL, did not influence the shape of standard curve. A parallelism test was carried out to compare the endogenous buffalo plasma prolactin with bovine prolactin standard. To validate the assay biologically, 11 Murrah buffaloes were given a third-generation antiprolactin (Norprolac; 10 mg/animal, i.m.). Blood samples were collected 1 d prior to the start of Norprolac administration and continued up to seventh day in an Ovsynch treatment program. In all animals, there were abrupt declines in prolactin concentrations following Norprolac treatments, which confirmed the biological validation of the EIA. After development and validation of EIA procedure, the concentration of plasma prolactin was determined efficiently in samples collected during both summer and winter samples.     

Using yellow rocket as a trap crop for diamondback moth (Lepidoptera: Plutellidae)

Yellow rocket, Barbarea vulgaris (R. Br.) variety arcuata, was evaluated as a trap crop for diamondback moth, Plutella xylostella (L.) (Lepidoptera: Plutellidae), in cabbage, Brassica oleracea L. variety capitata, in 2003 and 2004. In 2003, the numbers of P. xylostella larvae found in field plots of cabbage alone were 5.2-11.3 times higher than those on cabbage plants in plots that included cabbage and several rows of yellow rocket. In an outdoor experiment in screenhouses, P. xylostella oviposition on cabbage was compared among six treatments that varied in the percentage of yellow rocket in relation to cabbage (0, 4, 8, 16, 24, and 32% of the plants were yellow rocket). Results indicated that the percentage of eggs laid on cabbage decreased as the percentage of yellow rocket in the treatment increased, but this decrease was not significant beyond 20% of the plants being yellow rocket. In 2004, the numbers of P. xylostella larvae in field plots of cabbage alone were 1.6-2.4 and 1.7-2.8 times higher than numbers in treatments with 10 and 20% trap crop, respectively. Sticky trap and sweep net captures of P. xylostella adults indicated that within-field dispersal was reduced by the presence of yellow rocket and aggregation occurred around yellow rocket plants. Our study suggests that using yellow rocket as a trap crop may reduce P. xylostella infestations in cabbage fields, and this possibility is discussed in the context of general crop and insect pest management practices in crucifers.