Lower Hypocotyl Research Articles

Promotion of testa rupture during garden cress germination involves seed compartment-specific expression and activity of pectin methylesterases.

Pectin methylesterase (PME) controls the methylesterification status of pectins and thereby determines the biophysical properties of plant cell walls, which are important for tissue growth and weakening processes. We demonstrate here that tissue-specific and spatiotemporal alterations in cell wall pectin methylesterification occur during the germination of garden cress (Lepidium sativum). These cell wall changes are associated with characteristic expression patterns of PME genes and resultant enzyme activities in the key seed compartments CAP (micropylar endosperm) and RAD (radicle plus lower hypocotyl). Transcriptome and quantitative real-time reverse transcription-polymerase chain reaction analysis as well as PME enzyme activity measurements of separated seed compartments, including CAP and RAD, revealed distinct phases during germination. These were associated with hormonal and compartment-specific regulation of PME group 1, PME group 2, and PME inhibitor transcript expression and total PME activity. The regulatory patterns indicated a role for PME activity in testa rupture (TR). Consistent with a role for cell wall pectin methylesterification in TR, treatment of seeds with PME resulted in enhanced testa permeability and promoted TR. Mathematical modeling of transcript expression changes in germinating garden cress and Arabidopsis (Arabidopsis thaliana) seeds suggested that group 2 PMEs make a major contribution to the overall PME activity rather than acting as PME inhibitors. It is concluded that regulated changes in the degree of pectin methylesterification through CAP- and RAD-specific PME and PME inhibitor expression play a crucial role during Brassicaceae seed germination.

Defining the Site of Light Perception and Initiation of Phototropism in Arabidopsis

Phototropism is an adaptive response allowing plants to optimize photosynthetic light capture. This is achieved by asymmetric growth between the shaded and lit sides of the stimulated organ. In grass seedlings, the site of phototropin-mediated light perception is distinct from the site of bending; however, in dicotyledonous plants (e.g., Arabidopsis), spatial aspects of perception remain debatable. We use morphological studies and genetics to show that phototropism can occur in the absence of the root, lower hypocotyl, hypocotyl apex, and cotyledons. Tissue-specific expression of the phototropin1 (phot1) photoreceptor demonstrates that light sensing occurs in the upper hypocotyl and that expression of phot1 in the hypocotyl elongation zone is sufficient to enable a normal phototropic response. Moreover, we show that efficient phototropism occurs when phot1 is expressed from endodermal, cortical, or epidermal cells and that its local activation rapidly leads to a global response throughout the seedling. We propose that spatial aspects in the steps leading from light perception to growth reorientation during phototropism differ between grasses and dicots. These results are important to properly interpret genetic experiments and establish a model connecting light perception to the growth response, including cellular and morphological aspects.

HRS1 acts as a negative regulator of abscisic acid signaling to promote timely germination of Arabidopsis seeds.

In this work, we conducted functional analysis of Arabidopsis HRS1 gene in order to provide new insights into the mechanisms governing seed germination. Compared with wild type (WT) control, HRS1 knockout mutant (hrs1-1) exhibited significant germination delays on either normal medium or those supplemented with abscisic acid (ABA) or sodium chloride (NaCl), with the magnitude of the delay being substantially larger on the latter media. The hypersensitivity of hrs1-1 germination to ABA and NaCl required ABI3, ABI4 and ABI5, and was aggravated in the double mutant hrs1-1abi1-2 and triple mutant hrs1-1hab1-1abi1-2, indicating that HRS1 acts as a negative regulator of ABA signaling during seed germination. Consistent with this notion, HRS1 expression was found in the embryo axis, and was regulated both temporally and spatially, during seed germination. Further analysis showed that the delay of hrs1-1 germination under normal conditions was associated with reduction in the elongation of the cells located in the lower hypocotyl (LH) and transition zone (TZ) of embryo axis. Interestingly, the germination rate of hrs1-1 was more severely reduced by the inhibitor of cell elongation, and more significantly decreased by the suppressors of plasmalemma H+-ATPase activity, than that of WT control. The plasmalemma H+-ATPase activity in the germinating seeds of hrs1-1 was substantially lower than that exhibited by WT control, and fusicoccin, an activator of this pump, corrected the transient germination delay of hrs1-1. Together, our data suggest that HRS1 may be needed for suppressing ABA signaling in germinating embryo axis, which promotes the timely germination of Arabidopsis seeds probably by facilitating the proper function of plasmalemma H+-ATPase and the efficient elongation of LH and TZ cells.

Germination of Arabidopsis thaliana seeds is not completed as a result of elongation of the radicle but of the adjacent transition zone and lower hypocotyl

The completion of germination of seeds of Arabidopsis thaliana is marked by the appearance of the radicle through the surrounding endosperm and testa. Using confocal microscopy and green fluorescent protein (GFP)-transformed embryos to highlight the epidermal cell walls it has been possible to conduct time-lapse photography of individual embryos during their germination. This reveals that the elongation of embryo cells to effect completion of germination does not occur within the radicle itself, but rather within a discrete region that is immediately proximal to the radicle. This region, identifiable as the lower hypocotyl and hypocotyl-radicle transition zone, is also definable by accumulation of carbohydrate-containing bodies during germination, and distinct GFP expression of GAL4-GFP in enhancer trap lines. Flow cytometric studies show that there is an increase in the proportion of 4C nuclei in the axis which coincides with a considerable increase in length of the hypocotyl, and the occurrence of endopolyploid (8C and 16C) nuclei accompanies the 2-fold increase in mean cell size in the region of elongation, the lower hypocotyl, and hypocotyl-radicle transition zone. Thus the observed cell elongation during germination is accompanied by an increase in nuclear DNA content, and the resultant elongation of the axis to effect radicle emergence is due to cell expansion, not to cell division. When studying the molecular events involved in the completion of germination, therefore, it may be prudent to focus on this region of elongation.

더덕의 체세포배발생에서 전형층분화와 자엽수의 관계

Embryogenic callus was obtained from cotyledonary explants of Codonopsis lanceloata on Murashige & Skoog`s medium supplemented with 1 mg/L 2,4-D. Suspension cultures of the embryogenic calli were grown on a shaker at 100 strokes/min, and then the calli were subcultured for 2 weeks in 2,4-D-free medium to produce somatic embryo. In somatic embryos at the globular stage, cotyledon initials began to differentiate themselves in the near distal end of the procambial strand. Dicotyledons, tricotyledon, tetracotyledon and fused cotyledon were differentiated from the distal ends of two, three, four and circular procambial strands, respectively. Nearly circular procambial strand in lower hypocotyls were independently differentiated into two, three, four procambial tissues at cotyledonary node and cotyledons to form somatic embryos with dicotyledon, tricotyledon, tetracotyledon. If the distal subepidermal cells of globular embryo exclusively became cotyledon initials, the torpedo or cotyledonary embryo was characterized by somatic embryos with fused cotyledon.

Tissue position, explant orientation and naphthaleneacetic acid (NAA) affect initiation of somatic embryos and callus proliferation in Norway spruce (Picea abies)

Sections from mature zygotic embryos of Norway spruce exhibited different capacities for somatic embryo initiation. The upper hypocotyl part (Zone 2) was the most embryogenic, followed by the lower hypocotyl (Zone 3) and the apical zone (Zone 1); the root part (Zone 4) never initiated embryonal-suspensor masses (ESM). The embryogenic capacity of mature zygotic embryo is narrowly located in the vicinity of Zone 2. The frequency of embryos differentiating simultaneously ESM on Zones 1 and 3 is very low (0.6%) compared to those initiating ESM on Zones 1 and 2 (7%) or Zones 2 and 3 (16%). Elevated concentrations of naphthalene acetic acid (40 and 80 μM) reduced ESM initiation and callus proliferation on all sections but Zone 1. Highest initiation rate was obtained when explants were cultured with an apical-end-up orientation. Placing the explant basal-end-up partially inhibited the expression of embryogenic capacity, as well as decreasing the callus proliferation on Zone 3. A weak positive correlation (r=0.19, p < 0.001) was found between embryogenic capacity of the explant and proliferation rate of the derived callus.

Examination of morphological changes in the first formed protoxylem in Arabidopsis seedlings.

We examined morphological changes in the first-formed protoxylem vessels in Arabidopsis seedlings. Between 2.5 and 8 days after imbibition, mean hypocotyl and root length increased 1.52 and 23.3 times, respectively. In the 2.5-day-old seedlings, two continuous protoxylem vessels were present in the hypocotyl-root axis. In the 8-day-old upper hypocotyls, six protoxylem vessels were observed, and in the lower hypocotyls, four protoxylem vessels and one or two metaxylem vessels were observed. In the 8-day-old roots, there were two protoxylem vessels and two or three metaxylem vessels. Two protoxylem vessels in the hypocotyls connected to two metaxylem vessels in the roots of 8-day-old seedlings. At the 0.3-mm part below the hypocotyl-root boundary, the mean intervals of neighboring annular secondary wall thickenings in protoxylem vessels in 8-day-old roots were 12.9% larger than those in 2.5-day-old roots. In more apical parts of 8-day-old roots, the mean intervals fluctuated between 1.71 and 2.29 microm. In 8-day-old seedlings, metaxylem vessels were formed between 0.4 mm above the hypocotyl-root boundary and 17 mm below the boundary. The intervals in these regions were not extended so much as protoxylem vessels were collapsed. The first-formed protoxylem vessels presumably retain their water-conductive function after metaxylem formation.

A Monitoring System for Green Fluorescence Protein Gene-transformed Fusarium oxysporum in Melon Seedlings

Using melon seedlings at the cotyledon stage and genetically marked fungi, a system for monitoring pathogenic and nonpathogenic Fusarium oxysporum was devised in the present study. Protoplasts were prepared from three formae speciales (melonis, radicis-lycopersici and fragariae )of F. oxysporum and transformed with a synthetic gene for green fluorescence protein. Transformants were primarily isolated in the presence of hygromycin B and then screened by the emission of bright green fluorescence. Roots of melon seedlings were inoculated with fluorescing microconidia of these fungi, and fungal infection behavior was traced. Using fluorescence microscopy, we directly observed not only the fungus at the root surface, but also the mycelia elongating in the trachea of roots. Both pathogenic and nonpathogenic fungi germinated and hyphae elongated superficially on the surface of root. Only pathogenic fungi caused root necrosis at the inoculation site. Hyphae grew within the stem to induce constriction or cracking of lower hypocotyls, then causing wilting of the seedlings. Infection behavior of genetically marked pathogenic and nonpathogenic F. oxysporum could be successfully monitored after inoculation of cotyledons of seedlings.

Humic Acid Seed and Substrate Treatments Promote Seedling Root Development

Seed of Pelargonium ×hortorum L.H. Bailey `Freckles' (geranium) and Tagetes patula L. `Bonanza' (marigold) were soaked for 12, 24, or 48 h in solutions containing 0 (deionized water), 5000, 10,000, or 15,000 mg·L-1 humic acid (HA) or nutrient controls (NC) containing similar levels of nutrients prior to planting. Soaking in deionized water (DI) and NC treatments had no significant effect on root fresh weight. However, several of the HA treatments increased root fresh weight of marigold seedlings, and all increased geranium root fresh weight. Percentage of germination and shoot fresh weight were not significantly affected by treatment. Seed of Cucumis sativus L. `Salad Bush' (cucumber), Cucurbita pepo L. `Golden Summer Crookneck' (squash), `Freckles' geranium and `Bonanza' marigold were sown into 15-cell plug trays (5 mL volume), and the substrate was drenched with DI, 2500 or 5000 mg·L-1 HA, or 2500 or 5000 mg·L-1 NC. DI and NC treatments did not affect root fresh weight. However, cucumber, squash, and marigold seedlings germinated in substrate drenched with 2500 and 5000 mg·L-1 HA and geranium seedlings germinated in substrate drenched with 2500 mg·L-1 HA had significantly higher root fresh weight than did seedlings from all other treatments. Percentage of germination and shoot fresh weight were not significantly affected by treatment. `Salad Bush' cucumber and `Golden Summer Crookneck' squash seedlings germinated on germination towels soaked with 2500 or 5000 mg·L-1 HA, had significantly higher root fresh weight than did seedlings germinated on towels soaked with DI or NC solutions. Treatment with HA did not affect shoot fresh weight or the number of lateral roots. However, HA treatment increased the total length of lateral roots. The increase in lateral root growth occurred primarily in lateral roots developing from the lower hypocotyl.

300 UV-B INDUCED CHANGES IN PHENYLALANINE AMMONIALYASE (PAL) ACTIVITY AND UV-ABSORBING COMPOUNDS IN COTTON AND CUCUMBER SEEDLINGS

UV-B (UV) induced changes in PAL activity and UV-absorbing compounds were followed in cotton after 1 to 9 days and in cucumber after 1 to 14 days. UV increased PAL activity in the lower hypocotyl (LH) of cotton but had no effect on the upper portion. In general, PAL decreased with time, but UV treatment slowed that decline in the LH portion. Anthocyanin concentration declined with time in both portions. In cucumber cotyledons, UV had no effect on PAL. In cucumber leaves, there was no overall effect of UV; but there were significant interactions with time. In both cotyledons and in leaves, PAL decreased with time. As in LH cotton tissue, UV slowed the rate of decline of PAL in cucumber leaves. In leaves, UV absorbing compounds (at 330 nm) were increased by UV; in cotyledons, the increase in absorption was greater in controls than in UV-B irradiated seedlings. In cotton, changes in anthocyanins mirrored those in PAL, this was not the case for UV absorbing compounds in either species.

Extent and pattern of early soybean seedling colonization by Fusarium oxysporum and F. solani in naturally infested soil

Fusarium oxysporum colonized soybean [Glycine max (L.) Merrill cv. Essex] cotyledons 1 day after planting in a naturally infested soil and was present in 25% of tissue segments after 4 days. Both F. solani and F. oxysporum colonized the lower hypocotyl and emerging roots 2 and 3 days after planting, respectively. After 4 days, both fungi were found colonizing the elongating, upper portion of the hypocotyl, but the root tip remained free of the two fungi. The hypocotyl-root transition zone had the highest frequency of colonization (20–28% of 2 mm tissue segments) by each species at 4 days. F. solani had higher inoculum efficiencies for colonization than F. oxysporum. Tissue pasteurization tests indicated most early colonization is confined to the outer portions of tissues. The relative frequency and pattern of isolation of F. oxysporum and F. solani from lesions paralleled that of early colonization results, but only a small portion of hypocotyl and root colonization sites developed into lesions. These results support the hypothesis that non-pathogenic and pathogenic F. oxysporum and F. solani strains are primary and frequent colonizers of soybean tissues. Non-pathogenic isolates may have potential for biocontrol.

Aphanomyces raphani . [Descriptions of Fungi and Bacteria

Abstract A description is provided for Aphanomyces raphani . Information is included on the disease caused by the organism, its transmission, geographical distribution, and hosts. HOSTS: Species in the Cruciferae only. Brassica alboglabra, B. carnet, B. campestris var. pekinensis, B. chinensis, B. juncea, B. napobrassica, B. napus, B. nigra, B. oleracea (vars acephala, botrytis, capitata, gemmifera, gongylodes, italica, viridis), B. perviridis, B. rapa, B. robertiana, Crambe abyssinica, Eruca saliva, Lunaria annua , Raphanus raphanistrum, R. sativus, Sinapis alba . DISEASE: Black-root disease of radish; the fungus is a facultatively necrotrophic plant pathogen. Small black, water-soaked lesions on the lower hypocotyl and epidermis accompanied by slight longitudinal cracking of the subepidermal tissues become visible 20 days after sowing. In seedlings, the roots, hypocotyls, petioles and cotyledons become blackened and collapse. On older plants horizontal spread of the fungus produces characteristic constrictions and irregular bands around the circumference of the root (girdle disease). Subsequent penetration of the inner cortex results in blackening of the internal tissues in a portion or all of the root, accompanied by deformity of the roots and stunting of the tops due to retardation of growth. Lesions become invaded rapidly by secondary colonisers such as soft-rot bacteria. GEOGRAPHICAL DISTRIBUTION: Africa: South Africa. Asia: Japan. Australasia &amp; Oceania: Australia (NSW Qld), New Zealand. Europe: Germany, Rumania. North America: Canada (British Columbia, NS, Ont, Que), USA (California, IL IA, IN, ME to Florida, OR). South America: Venezuela. See CMI Distribution Maps of Plant Diseases 421. TRANSMISSION: In wet soil, by zoospores which accumulate at root tips in response to hypocotyl exudates or indole-3-aldehyde (59, 3937). The large number of crucifers (particularly cole crops) infected by A. raphani allows it to persist even in the absence of a radish crop. Oospores are generally believed to survive in soil, although there is no evidence to support this. The oospore germinates via a hypha which produces an apical sporangium.

Multiple Shoot Induction in vitro from the Hypocotyl of Germinating Embryos of Flax ( Linum usitatissimum L.)

Summary Growth responses have been studied for mature embryos of flax divided into two segments by a cut across the top of the hypocotyl and cultured on Monnier's basal medium with or without yeast extract (1.0 g 1-1) and BAP (0.05 mg 1-1). On basal medium without additives, adventive organ primordia were rapidly formed by the hypocotyl tissues adjacent to the cut: shoots on the lower segment and roots on the upper segment. BAP stimulated continuing production of greater numbers of shoot primordia on the lower hypocotyl, but prevented shoot elongation. Yeast extract retarded shoot and root growth, and also prevented the formation of friable wound callus on the lower cut surface of the hypocotyl. The effects of BAP and yeast extract together were additive. Somatic embryogenesis was not observed. Preliminary histological studies suggested the involvement of both epidermal and cortical cell layers in forming shoot primordia on the lower hypocotyl. Plants were readily regenerated from adventitious shoots formed on lower embryo segments. Segments cultured with BAP required transfer to a medium without growth regulators to allow shoot elongation before subculture.

Anatomy of Pregermination to Postgermination Embryos of Hibiscus cannabinus

Mature embryo, well advanced with two leaf primordia, bore different shapes of glandular trichomes on adaxial epidermis of cotyledons. The height and shape of the shoot apical dome changed from almost flat or concave to a conical one in successive plastochrons after germination. Zonation into single tunica, two metrameristem layers, flank meristem (almost radiating) and pith rib meristem, were distinct in the seedling shoot apex. Occurrence of just divided cells in the pith rib and flank meristem of the pregermination embryo was interesting. Cell activation increased considerably after germination. Minute mitochondria occurred all over the apical and cotyledonary cells. Root apex was of “closed type”. Procambia initiated in two loci, one near cotyledon base, the other near the subterminal part of root, phloem started at two loci (cotyledon and lower hypocotyl), xylem just below cotyledon base. Cotyledon bundles had discontinuous differentiation. Primordial bundles differentiated bidirectionally from base.

Factors Involved in the Opening of the Hypocotyl Hook of Cotton and Beans

Conditions influencing the opening of the bean (Phaseolus vulgaris L.) and cotton (Gossypium hirsutum L.) hypocotyl hook were defined. Such hooks were shown to undergo geotropic curvature; orientation of the hook with respect to gravity greatly affected the observed opening. Cotton and bean hooks behaved exactly opposite in regard to the presence of the cotyledons and apical bud. The cotton hook required the cotyledons for opening, but the corresponding tissue slowed or inhibited opening of the bean hook. With cotton, lower hypocotyl and root tissues stimulated hook opening, but with bean, the tissues below the hook section had little effect. Kinetin and gibberellic acid both modified hook opening in light and dark; the former was inhibitory and the latter was stimulatory. Indoleacetic acid, at concentrations above 10(-5) M, caused pronounced hook closing in red light but not in the dark. These effects were generally the same with both plants. In opening of the cotton hook, the cotyledons were not necessary as a light receptor tissue. None of the growth substances tested were able to substitute completely for the cotton cotyledon. Coumarin was a pronounced inhibitor of opening of the cotton hook, and this response was expressed to such a degree as to cause hook closure with bean tissue. Reduced oxygen levels inhibited hook opening in bean. Oxygen was required in processes subsequent to the light reaction, but not for the photochemical process.

The effect of 2,4-dichlorophenoxyacetic acid upon the metabolism and composition of soybean hypocotyl mitochondria

Dark-grown, 3-day-old soybean seedlings were sprayed with 1 mM 2,4-dichlorophenoxyacetic acid 24 hours before harvest. Mitochondria from 2,4-D-treated lower hypocotyls were found to be larger and showed greater incorporation in vivo, of amino acids into protein and phosphate into phospholipids and RNA, than mitochondria from untreated tissue. Mitochondria isolated from 2,4-D-treated hypocotyls showed an enhanced energy-dependent incorporation of amino acids into protein, although the incorporation of nucleoside triphosphates into the RNA of isolated mitochondria was not affected. No effect of 2,4-D, applied in vitro, was noted, and no enhancement of mitochondrial respiratory efficiency followed auxin treatment. A method of isolating mitochondria with a very low level of bacterial contamination is reported.

Indoleacetic Acid and Molecular Differentiation Evidence for Interaction

AbstractProteins of hypocotyls of bean were studied by electrophoresis. Proteins were extracted from hypocotyl segments of various stages of development starting with the relatively undifferentiated hook regions and proceeding by 2 cm segments down the hypocotyl. The proteins were the soluble (pH 7.4), the basic nuclear (histones), acidic ribonuclear and acidic chromosomal.Soluble proteins reflected differentiation of the hypocotyl in that lower hypocotyl segments had more different protein types than did the hook region. Indoleacetic acid (IAA) at 10−6M when applied to the lower hypocotyl appeared to induce still more different proteins. However, at 10−3M, IAA appeared to induce molecular dedifferentiation in that hypocotyl protein patterns began to resemble those of the hook.Histones also reflected differentiation, the hook having more histone types than the lower hypocotyl. IAA had no effect on histones.The hook region had two types of acidic chromosomal proteins, the lower hypocotyl one. When lower hypocotyl segments were incubated in 10−3M IAA, the protein pattern resembled that of the hook in that the second protein normally present in the hook and not in the hypocotyl was in fact induced in the hypocotyl.The hook had two acidic ribonuclear proteins, the lower hypocotyl one. IAA did not affect this protein.These experiments suggest that IAA in some manner regulates molecular (protein) differentiation. It is further suggested that IAA accomplishes this control through the acidic nuclear proteins which are closely associated with genetic material and which reflect differentiation and are also affected by IAA.

ROW TREATMENT OF SOIL WITH TETRAMETHYLTHIURAM DISULPHIDE FOR CONTROL OF BLACKROOT OF SUGAR-BEET SEEDLINGS. I. GREENHOUSE TESTS

The prevention of blackroot of sugar-beet seedlings has been attempted in greenhouse experiments, by row applications of different proprietary compounds to naturally-infested field soils representing three distinct types and differing biologically with respect to associated pathogens.Segmented seed, var. U.S. 215 × 216, was used in all the tests as was also a standard, commercial fertilizer. In earlier trials, the chemical under test and the fertilizer were added separately to a measured volume of rootrot soil. In later trials, the chemical mixed with the fertilizer was incorporated with the soil by a method approaching more closely that employed in field practice. The two procedures, designated as the precision and the surface application method, respectively, are described in detail. Calculations for the requisite amounts of chemical and fertilizer were made on the basis of sugar-beet rows in the field being spaced 22 in. apart, i.e., 23,760 linear feet of row per acre.Tetramethylthiuram disulphide (50%, nonwettable), when applied as Arasan or D-419 at the rate of 3 to 4 lb. per acre, consistently proved to be by far the most effective of the chemicals tested, in reducing pre- and postemergence black-root. Arasan, which was investigated more intensively than any of the other chemicals, was found (a) to retain its fungicidal capability after being mixed for as long as 14 months with a commercial fertilizer, (b) to be effective in reducing the incidence of the disease in three quite different types of soil, and (c) to remain effective in the soil over relatively long periods of time. These beneficial effects are contingent, however, upon correct placement of the chemical, best results having been obtained when the seed germinated in, and the seedlings grew up through, soil impregnated with the Arasan–fertilizer mixture. Up to 21 °C., Arasan, at a rate as high as 4 lb. per acre, will, in a moderately wet soil, effectually control the disease without injury to the seedlings. At 27° to 29 °C., moderate injury to roots and lower hypocotyl resulted from a 3 lb. per acre application of the chemical and, following a 4 lb. application, root and hypocotyl injury was accentuated, typical foliar symptoms appeared, and mortality of seedlings was appreciable.Of the other chemicals tested, 2,4,5-trichlorophenyl acetate (Mycotox No. 1), 20% copper trichlorophenate (C-119), 50% 2,4,5-trichlorophenyl chloroacetate (F-800), and a product of undisclosed chemical composition (CCH-358 or 358A) showed encouraging possibilities.

Vascular Anatomy of the Seedling of Melilotus alba

1. The primary root of Melilotus alba is characterized by a triarch radial protostele. 2. The first indication of a change from the triarch exarch arrangement of the xylem of the primary root in the young seedling is the development of diarchy or tetrarchy in the lower hypocotyl. 3. If tetrarchy occurs, it is invariably replaced at a higher level in the hypocotyl by diarchy. 4. The exarch ridge of xylem on the intercotyledonary axis gradually disappears at higher levels in the hypocotyl, and has no direct connection with the primary xylem of cotyledonary or plumular traces. 5. The exarch-endarch transition is continued in the formation of the cotyledonary traces and is completed in the main vascular bundles at the base of the cotyledonary blades. 6. A conspicuous pith differentiates in the hypocotyl, the stele enlarges, the cortex becomes more prominent, and a cutinized epidermis is present in this region in the young seedling. An endodermis with Casparian strips is present throughout the greater length of the hypocotyl. 7. A continuous, primary vascular system exists from root, through hypocotyl, into cotyledons. 8. The median vascular bundle which supplies the unifoliate leaf at the second node develops from meristematic tissue in a seedling several days old. The first indication of the differentiation of this collateral endarch bundle is the appearance of a single xylem element in the upper hypocotyl on the intercotyledonary axis, above the disappearing protoxylem of the third ray or on the opposite side. 9. In seedlings seven to ten days old, the median trace to the first trifoliate leaf at the third node, as well as the four bundles which continue upward to form the two lateral traces to the unifoliate leaf and the first trifoliate leaf, respectively, are well differentiated as endarch collateral bundles in the upper hypocotyl. These six primary bundles arise from meristematic tissue and appear to have no direct connection with the primary exarch strands in the lower hypocotyl. 10. Buds arise early in the axils of the cotyledons, the unifoliate leaf, and the first trifoliate leaf. The cotyledonary buds do not ordinarily develop into branches the first season. 11. Cambial activity occurs early in the primary root; fibers are prominent in the secondary xylem and phloem. As a result of cork cambium, which arises in the pericycle, a prominent cork is formed and the cortex of the root disappears. 12. The square stem is characterized by a dictyostele, with collateral endarch bundles. 13. The similarity between the method of transition from root to stem structure in Melilotus alba and the types described in M. arvensis, Medicago, Trigonella, Ononis, and Trifolium indicates a close structural and phylogenetic relationship between these species in the Trifolieae.