The Germination of Carrot (Daucus carota L.) Seed Harvested on Two Dates: A Physiological and Biochemical Study

The germination of two batches of carrot seed, harvested 44and 104 d after anthesis, was compared at 10 °C. Proteinand nucleic acid contents of the seeds were measured at intervals,together with fresh weight and respiration rate. The matureseed germinated 3.7 d earlier than the immature seed, with nodifference in percentage germination. The dry mature seed containedmore protein and nucleic acid per unit dry matter than did theimmature seed, and proportions of nucleic acid present as rRNAand poly(A)RNA were greater in the mature seed. The sequenceof metabolic reactivation was the same in both batches of seed,as were the relative rates of increase of each nucleic acidcomponent, fresh weight, and respiration rate. Differences inthe composition of the dry seed appear to be responsible forthe observed difference in rate of germination between the twoseed batches.     

Factors Influencing Embryogenesis in Carrot Cultures (Daucus carota L.)

There is no doubt that isolated vacuolated carrot cells cande-differentiate and give rise to embryogenic clusters fromwhich embryoids arise. However a study of the origins of embryogeniccells in culture suggests that the most frequent source is fromgroups of small meristematic cells liberated from the primaryexplant, and maintained as meristematic cells through numeroussubcultures. Groups of vacuolated cells divide to give riseto callus nodules, which can undergo morphogenesis in a varietyof ways. Transitions from one cell type to another are relativelyinfrequent, and cells generally divide to give rise to cellsof similar type to the parent. The occurrence of a low proportionof embryogenic cells in an inoculum is sufficient to resultin large numbers of embryoids when medium conditions are changedto favour their proliferation and development. The various routesby which plants can arise from carrot cultures are discussed.     

Inositol Trisphosphate Metabolism in Carrot (Daucus carota L.) Cells

The metabolism of exogenously added d-myo-[1-³H]inositol 1,4,5-trisphosphate (IP₃) has been examined in microsomal membrane and soluble fractions of carrot (Daucus carota L.) cells grown in suspension culture. When [³H]IP₃ was added to a microsomal membrane fraction, [³H]IP₂ was the primary metabolite consisting of approximately 83% of the total recovered [³H] by paper electrophoresis. [³H]IP was only 6% of the [³H] recovered, and 10% of the [³H]IP₃ was not further metabolized. In contrast, when [³H]IP₃ was added to the soluble fraction, approximately equal amounts of [³H]IP₂ and [³H]IP were recovered. Ca²⁺ (100 micromolar) tended to enhance IP₃ dephosphorylation but inhibited the IP₂ dephosphorylation in the soluble fraction by about 20%. MoO₄²⁻ (1 millimolar) inhibited the dephosphorylation of IP₃ by the microsomal fraction and the dephosphorylation of IP₂ by the soluble fraction. MoO₄²⁻, however, did not inhibit the dephosphorylation of IP₃ by the soluble fraction. Li⁺ (10 and 50 millimolar) had no effect on IP₃ metabolism in either the soluble or membrane fraction; however, Li⁺ (50 millimolar) inhibited IP₂ dephosphorylation in the soluble fraction about 25%.     

Characterization of Inositol Phosphates in Carrot (Daucus carota L.) Cells

We have shown previously that inositol-1,4,5-trisphosphate (IP₃) stimulates an efflux of ⁴⁵Ca²⁺ from fusogenic carrot protoplasts (M Rincón, WF Boss [1987] Plant Physiol 83: 395-398). In light of these results, we suggested that IP₃ might serve as a second messenger for the mobilization of intracellular Ca²⁺ in higher plant cells. To determine whether or not IP₃ and other inositol phosphates were present in the carrot cells, the cells were labeled with myo-[2-³H]inositol for 18 hours and extracted with ice-cold 10% trichloroacetic acid. The inositol metabolites were separated by anion exchange chromatography and by paper electrophoresis. We found that [³H]inositol metabolites coeluted with inositol bisphosphate (IP₂) and IP₃ when separated by anion exchange chromatography. However, we could not detect IP₂ or IP₃ when the inositol metabolites were analyzed by paper electrophoresis even though the polyphosphoinositides, which are the source of IP₂ and IP₃, were present in these cells. Thus, [³H] inositol metabolites other than IP₂ and IP₃ had coeluted on the anion exchange columns. The data indicate that either IP₃ is rapidly metabolized or that it is not present at a detectable level in the carrot cells.     

Cellular Basis of Tissue Toughness in Carrot (Daucus carota L.) Storage Roots

Carrots are brittle, tending to split and break on harvestingas a result of impacts. Since the phloem tissue of carrot storageroots is largely parenchymatous, there is little to resist thepropagation of a fracture once initiated. In carrots, fracturetoughness is inversely related to water potential, whereas tensilestrength is virtually independent of water status. Fracturetoughness tend to reflect changes in root water potential, buttensile strength increases on two separate occasions despitethe fact that root and cell expansion is accompanied by a reductionin apoplast volume fraction. Possible mechanisms to accountfor carrot tissue toughening towards the end of crop growthare discussed.Copyright 1995, 1999 Academic Press Daucus carota, carrot, toughness, strength, harvest damage, water status, apoplast volume fraction     

myo-Inositol Trisphosphate Mobilizes Calcium from Fusogenic Carrot (Daucus carota L.) Protoplasts

To determine whether or not inositol trisphosphate (IP₃) mobilizes calcium in higher plant cells, we investigated the effect of IP₃ on Ca²⁺ fluxes in fusogenic carrot (Daucus carota L.) protoplasts. The protoplasts were incubated in ⁴⁵Ca²⁺-containing medium and the ⁴⁵Ca²⁺ associated with the protoplasts was monitored with time. Addition of IP₃ (20 micromolar) caused a 17% net loss of the accumulated ⁴⁵Ca²⁺ within 4 minutes. There was a reuptake of ⁴⁵Ca²⁺ and the protoplasts recovered to their initial value by 10 minutes. Phytic acid (IP₆), also stimulated ⁴⁵Ca²⁺ efflux from the protoplasts. Both the IP_3⁻ and the IP_6⁻induced ⁴⁵Ca²⁺ efflux were inhibited by the calmodulin antagonist, trifluoperazine.     

Organic Acid Metabolism in Aluminum-Phosphate Utilizing Cells of Carrot (Daucus carota L.)

Carbohydrate metabolism in Al-phosphate utilizing cells of carrot[designated as IPG, Koyama et al. (1992) Plant Cell Physiol.33: 171], which grow normally in Al-phosphate medium accompaniedby citrate excretion, was investigated. The excretion of citratewas strongly related to the availability of sucrose in medium,indicating that citrate excretion was severely limited by sucrosein medium. The ratio of the amount of carbon in the excretedcitrate to the consumed sucrose, was significantly higher inIPG cells than in wild-type cells. When 50% of the sucrose inthe medium was consumed, the ratio was 0.6% for the IPG cellsand 0.2% the wild-type cells. Under these conditions, IPG cellsshowed altered citrate synthesis metabolism, which resultedin increased citrate production. Specific activity of mitochondrialcitrate synthase was higher in IPG cells than in wild-type cells,whereas the activity of cytosolic NADP-specific isocitrate dehydrogenasewas lower in IPG cells than in wild-type cells. (Received August 27, 1998; Accepted February 21, 1999)     

Smoke-saturated Water and Karrikinolide Modulate Germination,Growth, Photosynthesis and Nutritional Values of Carrot (Daucus carota L.)

Plant-derived smoke is a positive regulator of seed germination and growth with regard to many plant species. Of the several compounds present in plant-derived smoke, karrikinolide or KAR₁ (3-methyl-2H-furo[2,3-c]pyran-2-one) is considered to be the major active growth-promoting compound. To test the efficacy of smoke-saturated water (SSW) and KAR₁ on carrot (Daucus carota L.), two separate pot experiments were simultaneously conducted in the same environmental conditions. SSW and KAR₁ treatments were applied to the plants in the form of aqueous solutions of variable concentrations. Prior to sowing, seeds were soaked in the solutions of SSW (25.8 µg L⁻¹_, 51.6 µg L⁻¹,103.2 µg L⁻¹ and 258.0 µg L⁻¹) and KAR₁ (0.015 µg L⁻¹, 0.150 µg L⁻¹, 1.501 µg L⁻¹ and 15.013 µg L⁻¹). Percent seed germination, vegetative growth, photosynthesis and nutritional values were the major parameters through which the plant response to the applied treatments was investigated. The results obtained indicated a significant improvement in all the plant attributes studied. In general, SSW (51.6 µg L⁻¹) and KAR₁ (1.501 µg L⁻¹) proved optimum treatments for most the parameters. As compared to the control, 51.6 µg L⁻¹ of SSW and 1.501 µg L⁻¹ of KAR₁ increased the percent seed germination by 58.0% and 54.4%, respectively. Over the control, the values of plant height and net photosynthetic rate were enhanced by 33.9% and 40.9%, respectively, due to 51.6 µg L⁻¹ of SSW, while the values of these parameters were increased by 25.2% and 34.0%, respectively, due to 1.501 µg L⁻¹ of KAR₁. In comparison with the control, treatment 51.6 µg L⁻¹ of SSW increased the contents of β-carotene and ascorbic acid by 32.7% and 37.9%, respectively, while the treatment 1.501 µg L⁻¹ M of KAR₁ enhanced these contents by 42.0% and 48.4%, respectively. This study provides an insight into an affordable and feasible method of crop improvement.

     

The Effect of Cultivar and Cultural Factors on Embryo-sac Volume and Seed Weight in Carrot (Daucus carota L.)

The influence on carrot seed weight of cultivar, plant density,position of the seed on the plant, and of shading and removingumbels at different stages of growth was examined. CultivarDanvers produced seeds of approx. 2.5 mg, nearly twice as heavyas those of cv. Amsterdam with cv. Chantenay having an intermediateweight. Seeds from plants grown at 80 plants m^–2 were,on average, approx. 20 per cent lighter than those from plantsgrown at 10 plants m^–2 as were those from secondary thanprimary umbels. There were few differences in seed RGR betweencultivars, plant densities or seed positions and the differencesin seed weight produced by the treatments were largely associatedwith the duration of seed growth and with the volumes of theovary, ovule and embryo-sac at the time of fertilization. Shading(66 per cent reduction in irradiance) for the period of twoweeks before or two weeks after fertilization reduced seed weightcompared with unshaded plants. Removing the secondary and higherorder umbels from plants at these stages compared with no removalor removal at a later stage of growth increased seed weightof the remaining primary-umbel seed. Carrot, Daucus carota, seed weight, growth duration, RGR, embryo-sac, ovule, shading, pruning     

南鹤虱凝集素的研究

本文对伞形科16种植物(药用部分)是否存在凝集素进行了筛选。发现野胡萝卜果实与芹菜果实中有凝集素。野胡萝卜果实(中药名称南鹤虱)抽提液的凝血活性较芹菜强。 采用CM-纤维素及DEAE-纤维素对南鹤虱凝集素进行分离纯化。用聚丙烯酰胺凝胶电泳鉴定其纯度。用SDS-聚丙烯酰胺凝胶电泳测定亚基分子量为32600。 此凝集素对热及酸较稳定,对碱略差。对人的A、B、O血型无专一性。能凝集兎、小鼠、牛及鸡的血,但不凝集蟾蜍的血。 此凝集素的凝血活性能被D-木糖及N-乙酰葡萄糖胺轻微地抑制。 经shiff试剂检测表明南鹤虱凝集素为一糖蛋白。糖含量为3.4％。此凝集素分子中的谷氨酸门冬氨酸、精氨酸及絲氨酸的含量较高。     

Correction to: Smoke-saturated Water and Karrikinolide Modulate Germination,Growth, Photosynthesis and Nutritional Values of Carrot (Daucus carota L.)

Journal of Plant Growth Regulation - The original version of this article unfortunately contained a mistake in Table 3. The correct Table is given below.     

Transformation and regeneration of carrot (Daucus carota L.)

A protocol is presented for the efficient transformation of carrot (Daucus carota L. cv. Nantaise) by Agrobacterium tumefaciens. The binary vector contained the marker gene -glucuronidase (GUS), driven by the 35S promoter of cauliflower mosaic virus, and the nptII gene, which confers kanamycin resistance. Highest T-DNA transfer rates were obtained by co-cultivating bacteria with hypocotyl segments of dark-grown seedlings on solidified B5 medium containing naphthaleneacetic acid and 6-benzylaminopurine. After 2 days, bacterial growth was stopped with antibiotics. Two weeks later, the explants were placed on agar containing the kanamycin derivate geneticin; antibiotic-resistant calli developed during the following 4 weeks. Suspension cultures were obtained from resistant calli and plants regenerated via somatic embryogenesis in liquid culture. The majority of plants were phenotypically normal and, depending on the Agrobacterium strain used, harbored single or multiple copies of the T-DNA. About equal levels of GUS activity were found in different organs of young plants up to 6 weeks after embryogenesis. In leaves of older plants, GUS activity was markedly reduced, whereas the activities in phloem and xylem parenchyma cells of developing tap roots were still high and fairly uniform. Thus, the 35S promoter may be a useful tool to drive the expression of transgenes in developing carrot storage roots.     

Seed reserve-dependent growth responses to temperature and water potential in carrot (Daucus carota L.)

Both temperature and soil moisture vary greatly in the surface layers of the soil through which seedlings grow following germination. The work presented studied the impact of these environmental variables on post-germination carrot growth to nominal seedling emergence. The rapid pre-crook downward growth of both the hypocotyl and root was consistent with their requirement for establishment in soil drying from the surface. At all temperatures, both hypocotyl and root growth rates decreased as water stress increased and there was a very distinct temperature optimum that tended to occur at lower temperatures as water stress increased. A model based on the thermodynamics of reversible protein denaturation was adapted to include the effects of water potential in order to describe these growth rate responses. In general, the percentage of seedlings that reached the crook stage (start of upward hypocotyl growth) decreased at the extremes of the temperature range used and was progressively reduced by increasing water stress. A model was developed to describe this response based on the idea that each seedling within a population has lower and upper temperature thresholds and a water potential threshold which define the conditions within which it is able to grow. This threshold modelling approach which applies growth rates within a distribution of temperature and water potential thresholds could be used to simulate seedling growth by dividing time into suitable units.     

Root Development and Source-Sink Relations in Carrot, Daucus carota L.

Growth analysis and ¹⁴CO₂ feeding experiments have shown thatthe developing storage organ became an increasingly importantsink for assimilates, accumulating 40% of the dry matter producedby the carrot plant within 9 weeks of sowing. The relative importanceof each leaf in fixing and exporting ¹⁴C was assessed at twostages of development. Morphogenetic responses indicated thatan absence of thickening in the lateral roots was associatedwith continued meristematic activity in the tap root, in theform of an elongating apex or a vascular cambium. Source-sink relations were examined by observing plant growthfollowing the removal of part of the tap root and/or lateralroots. Pruning the roots at 35 d reduced the subsequent growthof the plant by reducing the AGR of the remaining root systemand the shoot. The reduction in leaf growth was associated witha loss of fibrous roots, removal of part of the tap root havingvery little additional effect on shoot growth although the AGRof the root system was reduced by a further 78%. Increased fibrousroot RGRs following pruning soon re-established the normal fibrousroot/shoot ratio. These experiments demonstrated the importanceof the root system in controlling dry matter production in thecarrot plant, but suggested that the sink activity of the developingstorage organ was less significant than other root functions.     

Surgical Experiments on the Differentiation of Vascular Tissue in the Shoot Apex of Carrot (Daucus carota L.)

Surgical techniques were applied to the shoot apex of carrot(Daucus carota L.) to test the interpretation that provasculartissue is the initial stage of vascular differentiation andto localize the sources of the influences that control its differentiation.If the apex is isolated laterally by vertical incisions leavingit at the summit of a plug of pith tissue, vascular differentiationproceeds normally and an independent vascular system is formedin the pith plug. If all leaf primordia are systematically suppressed,provascular tissue continues to differentiate as an acropetalextension of the pre-existing vascular system but no furtherdifferentiation occurs. When the apex is isolated laterallyand all leaf primordia are suppressed, provascular tissue continuesto be formed acropetally and is extended basipetally into thepith plug by redifferentiation of pith cells, but no furtherdifferentiation occurs. This tissue reacts positively to histochemicaltests for esterase indicating its vascular nature. If only oneleaf primordium is allowed to develop on an isolated shoot apex,its vascular system develops normally and extends basipetallyinto the pith plug, but there is no extension of provasculartissue into the pith plug. These results support the interpretationthat the initial stage of vascular differentiation is controlledby the apical meristem but that further maturation of vasculartissue depends upon influences from developing leaf primordia.Copyright 2000 Annals of Botany Company Provascular tissue, differentiation, carrot (Daucus carota L.), shoot apex, surgical techniques, leaf primordia     

Methylation of mitochondrial DNA in carrot (Daucus carota L.)

The methylation status of carrot (Daucus carota L.) mitochondrial DNA (mtDNA) was studied using isoschizomeric restriction enzymes MspI/HpaII (CCGG) and MvaI/EcoRII [CC(A/T)GG]. Southern hybridisations with probes for mitochondrial genes coxII and atpA were performed. MtDNAs isolated from non-embryogenic cell suspensions and roots were analysed. No differences were found using MspI/HpaII but after digesting the mtDNA with MvaI and EcoRII, some qualitative and quantitative differences between the restriction patterns appeared. Distinction was also revealed after Southern hybridisation with the coxII probe. These data indicate that the mtDNA of carrot is methylated in CNG trinucleotides and unmethylated in CG dinucleotides in CCGG sequences. The results were reproducible for cell suspensions of various genotypes and even cultivars but the extent of methylation was different in the root. The possible role of methylation in the mitochondrial genome of higher plants is discussed. Received: 16 April 1997 / Revision received: 4 July 1997 / Accepted: 30 July 1997