Asymbiotic seed germination and in vitro seedling development of Habenaria macroceratitis (Orchidaceae), a rare Florida terrestrial orchid

Continuing loss of native orchid habitat has lead to an increased emphasis on orchid conservation. Major obstacles in the production of native orchid seedlings for use in conservation have been: (1) development of efficient and reliable seed germination protocols and (2) an understanding of early seedling growth and development. Effects of six asymbiotic media (Modified Lucke, Murashige & Skoog, Lindemann, Vacin & Went, Malmgren Modified, Knudson C), four exogenous cytokinins (BA, Zea, Kin, 2-iP), and three photoperiods (0/24, 16/8, 24/0 h L/D) were examined on seed germination and early protocorm development of Habenaria macroceratitis, a rare native Florida terrestrial orchid. Finally, the effects of three photoperiods (8/16, 12/12, 16/8 h L/D) on in vitro seedling development were examined. Percent seed germination was highest on both LM and KC after seven weeks culture (LM = 89.1%, KC = 89.2%); however, protocorm development was enhanced on MM after both seven and 16 weeks. Both zeatin and kinetin at 1 μM enhanced seed germination (Zea = 58.1%, Kin = 47.2%). Final percent seed germination (91.7%) and protocorm development (Stage 4) was increased in the absence of light (0/24 h L/D). In␣vitro seedlings cultured under 8/16 h L/D conditions produced the highest number of tubers per seedling (1.06) with the greatest tuber (42.7 μg) and shoot (fwt = 69.5 μg) biomass and tuber diameter (3.1 mm).     

Azospirillum brasilense Sp245 triggers cytokinin signaling in root tips and improves biomass accumulation in Arabidopsis through canonical cytokinin receptors

The plant growth promoting rhizobacterium Azospirillum brasilense Sp245 enhances biomass production in cereals and horticultural species and is an interesting model to study the physiology of the phytostimulation program. Although auxin production by Azospirillum appears to be critical for root architectural readjustments, the role of cytokinins in the growth promoting effects of Azospirillum remains unclear. Here, Arabidopsis thaliana seedlings were co-cultivated in vitro with A. brasilense Sp245 to assess whether direct contact of roots with bacterial colonies or exposure to the bacterial volatiles using divided Petri plates would affect biomass production and root organogenesis. Both interaction types increased root and shoot fresh weight but had contrasting effects on primary root length, lateral root formation and root hair development. Cell proliferation in root meristems analyzed with the CYCB1;1::GUS reporter decreased over time with direct contact, but was augmented by plant exposure to volatiles. Noteworthy, the expression of the cytokinin-inducible reporters TCS::GFP and ARR5::GUS increased in root tips in response to bacterial contact, without being affected by the volatiles. In A. thaliana having single (cre1-12, ahk2-2, ahk3-3), double (cre1-12/ahk2-2, cre1-12/ahk3-3, ahk2-2/ahk3-3) or triple (cre1-12/ahk2-2/ahk3-3) mutations in canonical cytokinin receptors, only the triple mutant had a marked effect on plant growth in response to A. brasilense. These results show that different mechanisms are elicited by A. brasilense, which influence the cytokinin-signaling pathway.     

Long-distance ABA Signaling and Its Relation to Other Signaling Pathways in the Detection of Soil Drying and the Mediation of the Plant’s Response to Drought

In this article we review evidence for a variety of long-distance signaling pathways involving hormones and nutrient ions moving in the xylem sap. We argue that ABA has a central role to play, at least in root-to-shoot drought stress signaling and the regulation of functioning, growth, and development of plants in drying soil. We also stress the importance of changes in the pH of the leaf cell apoplast as influenced both by edaphic and climatic variation, as a regulator of shoot growth and functioning, and we show how changes in xylem and apoplastic pH can affect the way in which ABA regulates stomatal behavior and growth. The sensitivity to drought of the pH/ABA sensing and signaling mechanism is emphasized. This allows regulation of plant growth, development and functioning, and particularly shoot water status, as distinct from stress lesions in growth and other processes as a reaction to perturbations such as soil drying.     

Corticosterone responses of hand‐reared and parent‐reared grey‐faced petrel chicks (Pterodroma macroptera gouldi)

Hand‐reared grey‐faced petrel (Pterodroma macroptera gouldi) chicks (Order Procellariformes) that were subjected to a standardised blood sampling protocol immediately before they fledged showed a reduced corticosterone response compared to parent‐raised chicks. Serum corticosterone concentrations were lower in hand‐reared than parent‐reared birds 30 and 60 min after handling was initiated (21.5±6.7 vs. 105.4±7.4 ng/ml at 30 min, and 11.3±3.6 vs. 93.8±11.4 ng/ml at 60 min, respectively). The total integrated corticosterone response and the response corrected for initial corticosterone concentrations were similarly lower in hand‐reared than parent‐reared birds (927±489 vs. 4639±1924 ng/ml.min, and 529±352 vs. 4110±1896 ng/ml.min, respectively). Habituation to handling associated with hand‐rearing, which is expressed as a subdued corticostrone response, is likely to reduce the physiological stress associated with the translocation and reintroduction of these birds. However, the longer‐term consequences of this attenuated response remain unclear. Zoo Biol 0:1–8, 2005. © 2005 Wiley‐Liss, Inc.     

Plant development inhibitory genes in binary vector backbone improve quality event efficiency in soybean transformation

Conventional Agrobacterium-mediated plant transformation often produces a significant frequency of transgenic events containing vector backbone sequence, which is generally undesirable for biotechnology applications. We tested methods to reduce the frequency of transgenic plants containing vector backbone by incorporating genes into the backbone that inhibit the development of transgenic plants. Four backbone frequency reduction genes, bacterial levansucrase (sacB), maize cytokinin oxidase (CKX), Phaseolus GA 2-oxidase (GA 2-ox), and bacterial phytoene synthase (crtB), each expressed by the enhanced CaMV 35S promoter, were placed individually in a binary vector backbone near the left border (LB) of binary vectors. In transformed soybean plants, the lowest frequency of backbone presence was observed when the constitutively expressed CKX gene was used, followed by crtB. Higher backbone frequencies were found among the plants transformed with the GA 2-oxidase and sacB vectors. In some events, transfer of short backbone fragments appeared to be caused by LB readthrough and termination within the backbone reduction gene. To determine the effect of the backbone genes on transformation frequency, the crtB and CKX vectors were then compared to a control vector in soybean transformation experiments. The results revealed that there was no significant transformation frequency difference between the crtB and control vectors, but the CKX vector showed a significant transformation frequency decrease. Molecular analysis revealed that the frequency of transgenic plants containing one or two copies of the transgene and free of backbone was significantly increased by both the CKX and crtB backbone reduction vectors, indicating that there may be a correlation between transgene copy number and backbone frequency.     

植物的双组分信号系统

双组分系统由感受信号输入的组氨酸(His)蛋白激酶和调节信号输出的反应调控因子组成,涉及许多原核生物、真菌、黏菌和植物的各种信号转导途径.在植物中,还存在更复杂的包括杂合的His激酶、磷酸传递中间体和反应调控因子的信号系统,称为多步骤双组分系统.最近的研究表明,双组分系统在对环境刺激和生长调节剂(如乙烯、细胞分裂素、光和渗透胁迫)的反应中起重要作用.     

Benzyladenine Promotes Flowering in <Emphasis Type="Italic">Doritaenopsis</Emphasis> and <Emphasis Type="Italic">Phalaenopsis</Emphasis> Orchids

Two experiments were performed to determine how application of the cytokinin benzyladenine (BA) influenced flowering in Doritaenopsis and Phalaenopsis orchid clones. In the first experiment, two vegetative orchid clones growing in 15-cm pots were transferred from a 28°C greenhouse that inhibited flowering to a 23°C greenhouse for flower induction (day 0). A foliar spray (0.2 L m⁻²) containing BA at 100, 200, or 400 mg L⁻¹ or 25, 50, or 100 mg L⁻¹ each of BA and gibberellins A₄ + A₇ (BA+GA) was applied on days 0, 7, and 14. Plants treated with BA alone at 200 or 400 mg L⁻¹ had a visible inflorescence 3–9 days earlier and had a mean of 0.7–3.5 more inflorescences and 3–8 more flowers per plant than nontreated plants. The application of BA+GA had no effect on inflorescence number and total flower number at the rates tested. In the second experiment, three orchid clones received a single foliar spray of BA at 200 mg L⁻¹ at six time points relative to time of transfer from 29°C to 23°C (−1, 0, +1, +2, +4, or +6 weeks). A separate group of plants received a BA application at week 0 but was maintained at 29°C. Inflorescence number was greatest in all three orchid clones when plants were treated with BA 1 week after the temperature transfer. Plants that were sprayed with BA and maintained at 29°C did not initiate inflorescences. The promotion of flowering by the application of BA suggests that cytokinins at least partially regulate inflorescence initiation of Doritaenopsis and Phalaenopsis, but its promotion is conditional and BA application cannot completely substitute for an inductive low temperature.     

<Emphasis Type="Italic">Arabidopsis HOG1</Emphasis> gene and its petunia homolog <Emphasis Type="Italic">PETCBP</Emphasis> act as key regulators of yield parameters

Plant hormones influence the key parameters that contribute to crop yield, including biomass, branching and seed number. We tested manipulation of cytokinin signaling as an avenue for influencing these growth parameters. Here we report a full-length cDNA coding for a cytokinin binding protein, Petunia cytokinin binding protein (PETCBP) from Petunia hybrida cv. Mitchell. PETCBP encodes for a protein that exhibits high sequence similarity to S-adenosyl-L: -homocysteine hydrolase (SAHH). Transgenic petunia plants expressing this gene in antisense orientation displayed profuse branching, delayed flowering and delayed shoot bud induction from leaf explants in vitro. Homologs were also isolated from Arabidopsis thaliana homology-dependent gene silencing 1 (HOG1) and Orzya sativa (OsCBP). Arabidopsis HOG1 showed high affinity cytokinin binding activity and modified plant architecture similar to PETCBP. Transgenic Arabidopsis plants overexpressing HOG1 showed early flowering with a significantly reduced plant biomass and number of leaves. In contrast, profuse branching, delayed flowering, increased leaf size and higher seed yield were the major phenotypes observed in the antisense suppression lines. These results suggest that genetic manipulation of this cytokinin binding protein or its orthologs could be used for improving crop biomass and seed yield.     

Plant regeneration via protocorm-like body formation and shoot multiplication from seed-derived callus of a maudiae type slipper orchid

Tiny seeds from 5-month-old green capsules of a maudiae type slipper orchid, Paphiopedilum Alma Gavaert, were induced to form totipotent callus on 1/2 strength MS medium supplemented with 22.60 μM 2,4-D and 4.54 μM TDZ in darkness. The callus was proliferated more and maintained without any morphogenesis on the same medium with a 2-month interval of subculture for more than 2 years. When transferred to 1/2 MS medium supplemented with 26.85 μM NAA, an average of 4.7 protocorm-like bodies (PLBs)/shoot buds formed from each explant after 120 days of culture. After another 72 and 240 days of culture on the same medium, 25 shoot buds and eventually 75 plantlets were obtained through shoot multiplication from the original culture. Kinetin at 4.65 μM was suitable for shoot multiplication and could induce an average of 3.0 shoots from a single young shoot after 60 days of culture. The regenerated plantlets grew normally when transplanted to containers with sphagnum moss in a shaded greenhouse.     

Does habituation to humans influence predator discrimination in Gunther's dik-diks (Madoqua guentheri)?

Animals living around humans may habituate to us, but little is known about the consequences of this habituation. Some wildlife managers assume that habituation to humans makes individuals less likely to respond to natural predators, which is something to be avoided in captive breeding programmes where animals are destined for release. We conducted a playback experiment where we broadcast the sounds of a terrestrial predator and the song from a non-threatening bird to Gunther's dik-diks (Madoqua guentheri), a small ungulate that is vulnerable to many predators, in areas where dik-diks were and were not habituated to humans. Contrary to our expectation, habituated dik-diks discriminated the predator sounds from the birdsong, while unhabituated dik-diks failed to make this discrimination. Our results demonstrate that humans may influence predation hazard assessment, but we should not generally assume that human-habituated animals will be especially vulnerable to predators.