Seed germination,seedling inoculation and establishment ofAlnus spp. in containers in greenhouse trials

Methods for production of containerized seedlings ofAlnus species were developed which permit nitrogen-fixing nodules to form on the root systems prior to outplanting, in order to provide an early nitrogen input during seedling establishment. The methods are based on procedures for inoculating root systems with suspensions ofFrankia (Actinomycetales), applied either directly in the container cell as a soil drench at the time of seeding, or as a root dip for seedlings transplanted into the containers. Germination of dried, stored seed was enhanced by light and by presoaking for 16 h in water. Pretreatments to overcome seed dormancy or to eliminate fungal pathogens did not further enhance germination. Some loss of seedlings was recorded in the early stages of growth shortly after germination, which is a factor in calculating projected seedling yield. Nodulation and seedling growth were evaluated in terms of growth media characteristics. Seedlings performed well in peat-vermiculite, at soil pH between 5.5 and 8.0.     

APOLAR EMBRYOS OF FUCUS RESULTING FROM OSMOTIC AND CHEMICAL TREATMENT

Embryos of the brown alga Fucus vesiculosas L. were grown as populations in glass petri dishes in seawater at 15 C in continuous low-intensity unilateral fluorescent illumination for periods up to 2 weeks. A quantitative estimate of increase in nuclear number was made from acetocarmine squash preparations of samples taken at 12-or-24 hr intervals. Over the period of 2-6 days embryos showed a doubling time of about 12-18 hr. Under normal seawater culture conditions each embryo formed a single rhizoid. When grown in seawater supplemented with sugar concentrations above 0.4 m , Fucus embryos developed as multicellular spherical embryos lacking rhizoids. In 0.6 m sucrose-seawater, 97% of the embryos were apolar at 2 days; only 37% were apolar at 4 days, many having recovered from the sucrose inhibition. Some embryos remained apolar after growth in 0.6 m sucrose for 2 weeks. Nuclear counts showed that sucrose-seawater markedly inhibited the rate of cell division. Other sugars including D-glucose, D-fructose, D-galactose and the sugar alcohol D-mannitol were also effective. When apolar embryos grown in sucrose-seawater were returned to seawater, embryo growth resumed at the normal seawater rate, judged from nuclear counts. Such embryos formed multiple rhizoids, varying from two to eight rhizoids per embryo, which developed on the embryo quadrant or half away from the unilateral light. Each of the multiple rhizoids originated from a single small cell in the periphery of the multicellular spherica embryo. Thus the rhizoid-forming stimulus apparently had been subdivided among a number of the cells of the apolar embryos. The implications of this finding are discussed. Attempts to produce multiple rhizoids by treatment of embryos with indoleacetic acid or 2,4-dichlorophen-oxyacetic acid failed. However, embryos treated with 10⁻⁴ M or 5 × 10⁻⁵ m 2,3,5-triiodobenzoic acid formed 40 and 30% multiple rhizoids, respectively, suggesting that some chemical, perhaps hormonal, mechanism is involved in polarization and rhizoid initiation in Fucus embryogenesis.     

In vitro clonal multiplication of the actinorhizal plant Comptonia peregrina

Micropropagation of the actinorhizal plant Comptonia peregrina of the Myricaceae was achieved successfully by the induction of root buds in excised root culture with cytokinin (1.0 M benzyladenine). Excised root segments with initiated root buds were subcultured in Woody Plant Medium (WPM) lacking growth regulators, developing extensive callus which subsequently gave rise to multiple adventitious buds. Shoot elongation was facilitated by transfer of calluses to more aerated conditions. Root initiation was induced on shoots by brief treatment with auxin (<1 M indolebutyric acid) and transfer to WPM for plantlet development. Controlled light and aeration in liquid medium were critical conditions for successful micropropagation.     

Hormonal Control of Cell Proliferation and Xylem Differentiation in Cultured Tissues of Glycine max var. Biloxi

The relationship between tracheary element differentiation, cell proliferation and growth hormones was examined in agar-grown soybean callus. The time course of cell division and tracheary element formation in tissues grown on a medium containing 5 x 10(-7)m kinetin and 10(-5)m NAA was determined by means of maceration technique. After a slight lag period, a logarithmic increase in cell number was observed through the twelfth day of the culture period. Cell numbers increased at a considerably slower rate after the twelfth day. The rate of tracheary element formation varied with the rate of cell proliferation. Tracheary elements increased logarithmically during the log phase of growth. As the rate of cell division decreased after the twelfth day of culture, the rate of tracheary element formation also decreased. In the presence of 10(-5)m NAA, cell number increased as the kinetin concentration was increased between 10(-9) and 10(-6)m. However, tracheary element formation was not initiated unless the kinetin concentration was 5 x 10(-8)m or above. When the Biloxi callus was subcultured repeatedly on media containing 10(-8)m kinetin, a tracheary element-free population of cells was obtained. This undifferentiated tissue produced tracheary elements upon transfer to a medium containing 5 x 10(-7)m kinetin. In the presence of 5 x 10(-7)m kinetin, NAA stimulated cell proliferation between 10(-7) and 10(-5)m, but no tracheary elements were formed without auxin, or with 10(-7)m NAA. Neither NAA nor kinetin at any concentration tested stimulated tracheary element formation in the absence of an effective level of the other hormone. However, 2,4-D at 10(-7) or 10(-6)m promoted both cell proliferation and tracheary element differentiation in the absence of an exogenous cytokinin.     

Amino Acid incorporation in developing fucus embryos

The incorporation of ¹⁴C-leucine and ¹⁴C-amino acid mixture into protein in unfertilized eggs and developing embryos of the brown alga Fucus vesiculosus L. was studied. Bacterial contamination was initially a problem, but it was found that the addition of 40 μg/ml chloramphenicol to the incubation medium would inhibit bacterial protein synthesis without affecting early development of the Fucus embryos. The kinetics of uptake and incorporation of ¹⁴C-leucine into the trichloroacetic acid-soluble and -insoluble fractions indicated that the exogenous precursor did not equilibrate with the main soluble leucine pool before incorporation into protein. Uptake and incorporation of leucine by embryos 90 to 175 minutes old were proportional to exogenous leucine concentration over the range 5 × 10⁻⁶ m to 5 × 10⁻³ m. Unfertilized eggs will incorporate ¹⁴C-leucine into protein. The rate of this incorporation increases dramatically in newly fertilized eggs with a maximum rate at 3.5 hours, a period of cell wall formation and increasing metabolic rates. Thereafter, the rate of incorporation declines until approximately 15 to 17 hours when it increases again concurrently with the onset of rhizoid initiation and cell division.     

Phylogenetic diversification of immunoglobulin genes and the antibody repertoire

Immunoglobulins are encoded by a large multigene system that undergoes somatic rearrangement and additional genetic change during the development of immunoglobulin-producing cells. Inducible antibody and antibody-like responses are found in all vertebrates. However, immunoglobulin possessing disulfide-bonded heavy and light chains and domain-type organization has been described only in representatives of the jawed vertebrates. High degrees of nucleotide and predicted amino acid sequence identity are evident when the segmental elements that constitute the immunoglobulin gene loci in phylogenetically divergent vertebrates are compared. However, the organization of gene loci and the manner in which the independent elements recombine (and diversify) vary markedly among different taxa. One striking pattern of gene organization is the "cluster type" that appears to be restricted to the chondrichthyes (cartilaginous fishes) and limits segmental rearrangement to closely linked elements. This type of gene organization is associated with both heavy- and light-chain gene loci. In some cases, the clusters are "joined" or "partially joined" in the germ line, in effect predetermining or partially predetermining, respectively, the encoded specificities (the assumption being that these are expressed) of the individual loci. By relating the sequences of transcribed gene products to their respective germ-line genes, it is evident that, in some cases, joined-type genes are expressed. This raises a question about the existence and/or nature of allelic exclusion in these species. The extensive variation in gene organization found throughout the vertebrate species may relate directly to the role of intersegmental (V<==>D<==>J) distances in the commitment of the individual antibody-producing cell to a particular genetic specificity. Thus, the evolution of this locus, perhaps more so than that of others, may reflect the interrelationships between genetic organization and function.      

Early development ofRhizobium-induced root nodules ofParasponia rigida. I. Infection and early nodule initiation

Summary The first of two major steps in the infection process in roots ofParasponia rigida (Ulmaceae) following inoculation byRhizobium strain RP501 involves the invasion ofRhizobium into the intercellular space system of the root cortex. The earliest sign of root nodule initiation is the presence of clumps of multicellular root hairs (MCRH), a response apparently unique amongRhizobium-root associations. At the same time or shortly after MCRH are first visible, cell divisions are initiated in the outer root cortex of the host plant, always subjacent to the MCRH. No infection threads were observed in root hairs or cortical cells in early stages. Rhizobial entry through the epidermis and into the root cortex was shown to occur via intercellular invasion at the bases of MCRH. The second major step in the infection process is the actual infectionper se of host cells by the rhizobia and formation of typical intracellular infection threads with host cell accommodation. This infection step is probably the beginning of the truly symbiotic relationship in these nodules. Rhizobial invasion and infection are accompanied by host cortical cell divisions which result in a callus-like mass of cortical cells. In addition to infection thread formation in some of these host cortical cells, another type of rhizobial proliferation was observed in which large accumulations of rhizobia in intercellular spaces are associated with host cell wall distortion, deposition of electron-dense material in the walls, and occasional deleterious effects on host cell cytoplasm.