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Daniel I. Leskovar, Virgil Esensee, and Helen Belefant-Miller

Spinach (Spinacia oleracea L.) seed germination can be inhibited by high temperatures. An understanding of thermoinhibition in spinach is critical in predicting germination and emergence events. The purpose of this study was 3-fold: 1) to determine seed germination percentage and rate of spinach genotypes—`Cascade', `ACX 5044', `Fall Green', and `ARK 88-354'—exposed to constant and alternating temperatures; 2) to determine the nature and extent of inhibition imposed by the pericarp; and 3) to investigate leachate and oligosaccharide involvement in thermoinhibition. Germination inhibition began at >20 °C constant temperature and was totally suppressed at 35 °C. Alternating temperatures at 30/15 °C (12-hour day/12-hour night) resulted in greater germination than a constant 30 °C. The genotype sensitivity to supraoptimal temperatures was in the order of `ARK 88-354' ≤ `Fall Green' < `ACX 5044' < `Cascade', but the highly thermoinhibited `Cascade' seeds retained the ability to germinate when shifted to lower incubation temperatures. The pericarp inhibited germination, since seeds deprived of the pericarp had ≈90% germination at 30 °C. `ACX 5044' and `Cascade' had higher ABA content in the pericarp than `ARK 88-354' and `Fall Green'. Before imbibition at 30 °C, raffinose levels in each genotype were in the order of `ARK 88-354' > `Fall Green' > `Cascade'. After 48 hours of imbibition, sucrose and glucose levels were highest and raffinose levels were lowest in `ARK 88-354' and `Fall Green' seeds, while `Cascade' seeds remained less active metabolically. These data suggest that the pericarp apparently acts as a physical barrier as well as a source of inhibitors during thermoinhibition.

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Carlos A. Parera, Daniel J. Cantliffe, D.R. McCarty, and L. Curtis Hannah

The shrunken-2 (sh2) mutant of maize (Zea mays L.) increases sucrose and reduces starch in developing endosperm. An associated trait is poor seed and seedling vigor in seeds containing the mutation. The specific effects of sh2 mutant endosperm on embryo and seedling vigor were determined by analyzing seeds that contained either concordant wild-type or nonconcordant combinations of mutant and wild-type embryo and endosperm genotypes. The nonconcordant seed types that contained a wild-type embryo in association with a sh2 mutant endosperm or a sh2 mutant embryo in association with a wild-type endosperm were generated using the TB-3La translocation chromosome in which a wild-type Sh2 gene is attached to the centromeric portion of a B chromosome. Under stress conditions (complex stress vigor test), the seeds with mutant endosperm had lower germination, seedling fresh and dry weight, and index of conductivity than seeds with wild-type endosperm. Mutant endosperm and embryos excised from mutant endosperm imbibed more water than wild-type endosperm or embryos excised from wild-type endosperm. Because of the high concentration of osmotic solutes in the mutant endosperm, a rapid water uptake may induce a membrane disorganization. Leachate conductivities of seeds with mutant endosperm were higher than seeds with wild-type endosperm. In addition, a higher sucrose content and a lower raffinose to sucrose ratio were measured in the wild-type embryos associated with mutant endosperms than in the normal embryos excised from concordant wild-type seeds. These results suggest that a high rate of water uptake caused by the elevated concentration of osmotic solutes in seeds with mutant endosperms may affect membrane integrity during imbibition. Alternatively, the lower raffinose to sucrose ratio present in the mutant endosperm class might affect stabilization of cell membranes during seed desiccation. Embryos cultured in media containing 10% starch or no carbohydrate produced smaller seedlings than embryos cultured in 5% or 10% sucrose. Wild-type embryos excised from mutant endosperms exhibited lower germination in 0% and 5% sucrose media than embryos from concordant seed, indicating that reduced water uptake rates associated with lower external osmotic potential (10% sucrose) can improve vigor of embryos associated with sh2 mutant endosperm. The reduced vigor of embryos and seedlings that develop in association with sh2 mutant endosperm can be traced to the physiological and biochemical effects of the elevated sucrose levels present during seed formation and imbibition.

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Seenivasan Natarajan and Jeff Kuehny

Small heat shock proteins (sHSP) are a specific group of highly conserved proteins produced in almost all living organisms under heat stress. These sHSP have been shown to help prevent damage at the biomolecular level in plants. One of the greatest impediments to production of marketable herbaceous plants and their longevity is high temperature stress. The objectives of this experiment were to study the plant responses in terms of sHSP synthesis, single leaf net photosynthesis, total water-soluble carbohydrates (WSC), and overall growth for two S. splendens cultivars differing in performance under heat stress. `Vista Red' (heat tolerant) and `Sizzler Red' (heat sensitive) were exposed to short duration (3 hours) high temperature stresses of 30, 35, and 40 °C in growth chambers. Increasing the temperature to about 10 to 15 °C above the optimal growth temperature (25 °C, control) induced the synthesis of sHSP 27 in S. splendens. Expression of these proteins was significantly greater in the heat-tolerant vs. the heat-sensitive cultivar. Soluble carbohydrate content was greater in `Vista Red', and in both the cultivars raffinose was the primary soluble carbohydrate in heat-stressed plants. Overall growth of plants was significantly different in the two cultivars studied in terms of plant height, stem thickness, number of days to flower, and marketable quality. The better performance of `Vista Red' under heat stress was attributed to its morphological characteristics, including short stature, thicker stems and leaves. sHSPs and WSC are also found to be associated with heat tolerance and heat adaptation in S. splendens.

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Peter R. Hicklenton and Julia Y. Reekie

In northern regions, strawberry nursery plants are often dug in the late fall, packed and stored for winter, and shipped to markets in the early spring. Success depends on identifying when plants are dormant and can be safely stored. Beginning on 11 Oct., plants of Kent and Veestar were dug at weekly intervals from three fields in the Annapolis Valley, N.S., Canada. At each digging date root respiration was measured at 5, 10, 20 and 30°C. Six “first daughter” plants of each cultivar were wrapped in plastic and placed in ≈1.5°C refrigerated storage. Other plants were separated into roots and leaves for carbohydrate analysis. Fall temperatures were relatively mild with 417 crown chilling hours (8°C base) accumulated to 7 Nov. Only those plants dug on 11 Oct. did not survive when planted to the field on 1, June but vigor (number of daughters/runners) improved for plants dug later in the fall. For Kent, vigor increased through the last digging date (5 Dec.), but for Veestar, vigor did not change after 7 Nov. Early dug plants had relatively high rates of root respiration, low concentrations of leaf and root glucose, fructose, sucrose, and raffinose and high leaf starch, and low root starch concentrations. Most leaf sugar concentrations increased rapidly after 7 Nov., and root starch reached a maximum at the same date. Leaf and root carbohydrate concentrations were correlated with poststorage field vigor and may reflect the degree of plant dormancy at time of digging.

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Natalie L. Hubbard, D. Mason Pharr, and Steven C. Huber

Muskmelon (Cucumis melo L.) fruit lack a stored starch reserve and therefore depend on translocated photoassimilate from the leaf canopy for sugar accumulation during ripening. The influence of canopy photosynthesis on sucrose' accumulation within muskmelon fruit mesocarp was examined. Canopy photosynthetic activities were estimated in a sweet and a nonsweet genotype. Photosynthetic rate of the nonsweet genotype, on a per-plant basis, was only 56% of that of the sweet genotype. The effect of limiting leaf area of the sweet genotype on carbohydrate concentrations and sucrose metabolizing enzymes within the fruit was evaluated. A 50% reduction of leaf area 8 days before initiation of fruit sucrose accumulation resulted in canopy photosynthesis similar to that of the nonsweet genotype. Reduced photosynthetic activity resulted in slightly lower soluble-carbohydrate concentration in the fruit; however, fruit sucrose concentration was three times higher than that reported previously for the nonsweet genotype. The extent to which `fruit sucrose phosphate synthase (SPS) activity increased during maturation was diminished by leaf removal. Acid invertase activity declined in all fruit in a similar manner irrespective of defoliation. A reduction of leaf area of a sweet genotype reduced sucrose accumulation within the fruit. Lower fruit sucrose concentration was associated with lower concentration of raffinose saccharides and lower SPS activity within the fruit. Additionally, insufficient assimilate supply was judged not to be the factor responsible for low sucrose accumulation in a nonsweet genotype.

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Brian A. Birrenkott, Cynthia A. Henson, and Elden J. Stang

Cranberry (Vaccinium macrocarpon Ait. cv. Searles) vegetative tissue was analyzed at various stages of development to determine carbohydrate levels under field and greenhouse conditions and to identify the carbohydrates. Except during dormancy, cranberry uprights in the field had the highest concentration of carbohydrates (soluble and starch) at early blossom, when the lower flowers were at anthesis. As early flowers developed into fruit and upper flowers were at or just beyond anthesis, uprights had lower carbohydrate concentrations. As fruit growth slowed, soluble carbohydrate levels increased and were highest at dormancy. Upright shoot tissue produced the previous year and trailing woody stems followed the same trend as the current season's growth but had consistently lower soluble carbohydrate levels at each growth stage. Starch levels were low in current growth and did not change appreciably with fruit development. Starch was primarily stored and subsequently depleted in the previous season's upright growth and trailing woody stems. Tissue from the greenhouse was generally higher in carbohydrates than was field-grown tissue. Fruit developed from 53% of the flowers under greenhouse conditions, compared to 38% in the field. Insufficient carbohydrate levels may be responsible for the low fruit set observed in the field. Sucrose, glucose, fructose, raffinose, and stachyose were present in cranberry vegetative tissue.

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Abdul K. Janoudi, Irvin E. Widders, and James A. Flore

Cucumber plants were cultured in a greenhouse and subjected to either well-watered or water deficit conditions that reduced leaf water potential to-0.6 MPa. Leaf gas exchange measurements were conducted using an open gas exchange system. Carbon dioxide assimilation (A) attained saturation at a photon flux density (PFD) of 1000 μmol·m-2·s-1 (400-700 nm). There were no significant differences in A at ambient temperatures between 16 and 34C. Water use efficiency decreased rapidly with increasing vapor-pressure deficits to 2.5 kPa. Water stressed plants had lower stomata1 conductances and CO2 assimilation rates. The decrease in A was only partially due to stomata1 closure. The A vs. intercellular CO (Ci) relationship for stressed leaves revealed a change in the CO, compensation point, and that nonstomatal factors were contributing to the decrease in A in stressed plants. Thus, feedback inhibition of A may have occurred through photoassimilate accumulation. The concentrations of sucrose and raffinose were higher, and the concentration of stachyose was lower in leaves of stressed than of well-watered plants.

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D.B. Rowe, F.A. Blazich, D.M. Pharr, and F.C. Wise

Containerized, 2.5-year-old, hedged stock plants of four, full-sib families of loblolly pine (Pinus taeda L.) were fertilized daily with a complete nutrient solution containing 10, 25, 40, 55, or 70 ppm N, which resulted in a range of stock plant soluble carbohydrate (SCHO) and tissue N levels. SCHOs included myo-inositol, glucose, fructose, sucrose, and raffinose. Nitrogen concentrations and SCHO: N ratios ranged from 1.23% to 2.24% and 16:1 to 29:1, respectively. Softwood cuttings were taken in May and July 1995 and placed under intermittent mist. May cuttings rooted at significantly greater percentages than July cuttings (60% vs. 34%). Averaged over all N treatments, the best rooting family (56%) contained the highest tissue concentration of SCHOs (465 mg·g–1 dry weight) and had the highest SCHO: N ratio (26:1), whereas, the poorest rooting family (39%), exhibited the lowest level of SCHOs (357 mg·g–1 dry weight) and the lowest SCHO: N ratio (21:1). Rooting exhibited a quadratic response in regards to N fertilization levels and tissue SCHO concentrations. For both rooting trials, maximum rooting (83%) was noted for May cuttings taken from stock plants of one family fertilized with 40 ppm N, which corresponded to a tissue N concentration of 1.95% and a SCHO: N ratio of 22:1.

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Emma L. Locke*, Cecil Stushnoff, Joyce C. Pennycooke, and Michelle Jones

Salinity, drought and temperature frequently limit crop productivity. Transgenic Petunia ×hybrida cv. Mitchell with altered endogenous raffinose family oligosaccharides (RFO) due to over-expression (sense) or under-expression (antisense) of the tomato α-galactosidase gene show that antisense increases in RFO are associated with greater tolerance to freezing stress (Pennycooke et al., 2003). Because vegetative propagules of these antisense lines rooted and established more quickly than their sense counterparts, we hypothesized that antisense lines would also respond to salinity and wilting stress. Salinity treatment plants were exposed to 50-200 mm NaCl graduated 25 mm every 3 days and held at 200 mm for 13 days. Dry-down treatments were watered to pot capacity, then not watered until the onset of wilting. This was repeated in cycles for 26 days. Data were collected on plant growth, root/shoot ratios, and leaf water potential. Fresh and dry weights in four of the six antisense lines exceeded the wild type and sense lines. Osmotic potential for salinity and dry-down plants was 160% to 220% higher than control plants. Pearson correlations revealed that higher osmotic potential was partially associated with higher fresh weight (r = 0.7214, P = 0.02) and root/shoot ratios (r = -0.7414, P = 0.02) in salinity stressed plants. In the dry-down drought stressed plants, osmotic potential was not associated with fresh weight (r = 0.3364, ns) nor root/shoot ratio (r = -0.0431, ns). Salinity stress reduced root mass compared to control and dry down plants. Sense plants grew slowly and were highly variable.

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Manfredo J. Seufferheld, Cecil Stushnoff, John Fitzpatrick, and Philip L. Forsline

Cryopreservation of woody-plant, dormant buds may provide cost-effective, long-term, back-up conservation of germplasm for vegetatively propagated crops that are presently maintained as trees in field gene banks. Dormant buds can be recovered quickly by grafting to dwarfing rootstocks, thus producing flowers for breeding purposes, with minimum potential for inducing somaclonal variants. These attributes are essential to preserve the clonal integrity of unique gene combinations such as those found in tree fruit crops. Previous research has shown that dormant buds from cold-hardy apples can be recovered from storage in liquid nitrogen (LN) with high survival rates (80% to 100%) using controlled desiccation and slow freezing before immersing in LN. On the other hand, dormant buds from cold-tender taxa and buds collected at less than optimal stages for desiccation and freezing have much lower (0% to 50%) survival rates. We increased survival of cold-tender taxa by using a modified vitrification procedure. Dormant apple buds from tender and hardy cultivars were perfused with modified PVS [15% (w/v) ethylene glycol, 15% (w/v) propylene glycol, 7% (w/v) DMSO, and 15% (w/v) glycerol in 0.5 m sorbitol]. Toxicity from the PVS was reduced by dilution soaking in 1 m sorbitol, 0.2 m raffinose, and 15 mm CaCl2 before and after quench-freezing and slow-freezing cryopreservation. Up to 100% of some cold-tender taxa survived. In addition, xylem ray parenchyma tissues that supercool and are normally killed at about -40C with the desiccation protocol survived this vitrification procedure.