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  • 1
    ISSN: 1617-4623
    Keywords: Acremonium sp. ; Transformation ; Chromosomal karyotype ; Surrogate grass transformation ; uidA andhph genes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Conditions have been developed for transforming protoplasts of the perennial ryegrass endophyteAcremonium strain 187BB. Unlike most other ryegrass endophytes, this strain does not produce the lolitrem B neurotoxin and is therefore suitable as a host for surrogate introduction of foreign genes into grasses. Transformation frequencies of 700–800 transformants/μg DNA were obtained for both linear and circular forms of pAN7-1, a hygromycin (hph) resistant plasmid. Up to 80% of the linear transformants were stable on further culturing but only 25% of the circular transformants retained hygromycin resistance. Integration of pAN7-1 into the genome was confirmed by Southern blotting and probing of genomic digests of transformant DNA. Both single and tandemly repeated copies of the plasmid were found in the genome and both the number and sites of integration varied among the transformants. At least 13 chromosomes were identified in 187BB using contour-clamped homogeneous electric field (CHEF) gel electrophoresis. Probing of Southern blots of these gels confirmed that pAN7-1 had integrated into different chromosomes. The β-glucuronidase (GUS) gene,uidA, was also introduced into 187BB by co-transformation of pNOM-2 with pAN7-1. GUS activity was detected by growing the transformants on plates containing 5-bromo-4-chloro-3-indolyl β-D-glucuronic acid and by enzyme assays of mycelial extracts. Severalhph- anduidA-containing transformants were reintroduced into ryegrass seedlings and expression of GUS visualized in vivo, demonstrating that 187BB can be used as a surrogate host to introduce foreign genes into perennial ryegrass. Molecular analysis of fungal isolates from the leaf sheath confirmed that the pattern of pAN7-1 and pNOM-2 hybridizing fragments was identical to that observed in the fungus used as inoculum.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1572-9788
    Keywords: disease resistance ; glucose oxidase ; Gossypium hirsutum ; Nicotiana tabacum ; Talaromyces flavus ; transgenic plant
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Glucose oxidase secreted by the fungus Talaromyces flavus generates, in the presence of glucose, hydrogen peroxide that is toxic to phytopathogenic fungi responsible for economically important diseases in many crops. A glucose oxidase gene from T. flavus, was modified with a carrot extensin signal peptide and fused to either a constitutive or root-specific plant promoter. T1 tobacco plants expressing the enzyme constitutively were protected against infection by the seedling pathogen Rhizoctonia solani. Constitutive expression in tobacco was associated with reduced root growth, and slow germination on culture medium, and with reduced seed set in glasshouse conditions. Several independent transformed cotton plants with a root-specific construct expressed high glucose oxidase activity in the roots, excluding the root tip. Selected T3 homozygous lines showed some protection against the root pathogen, Verticillium dahliae, but not against Fusarium oxysporum. High levels of glucose oxidase expression in cotton roots were associated with reduced height, seed set and seedling germination and reduced lateral root formation. If this gene is to be of value for crop protection against pathogens it will require precise control of its expression to remove the deleterious phenotypes.
    Type of Medium: Electronic Resource
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  • 3
    Publication Date: 2019-10-23
    Description: Knowledge on basic biological functions of organisms is essential to understand not only the role they play in the ecosystems but also to manage and protect their populations. The study of biological processes, such as growth, reproduction and physiology, which can be approached in situ or by collecting specimens and rearing them in aquaria, is particularly challenging for deep-sea organisms like cold-water corals. Field experimental work and monitoring of deep-sea populations is still a chimera. Only a handful of research institutes or companies has been able to install in situ marine observatories in the Mediterranean Sea or elsewhere, which facilitate a continuous monitoring of deep-sea ecosystems. Hence, today’s best way to obtain basic biological information on these organisms is (1) working with collected samples and analysing them post-mortem and / or (2) cultivating corals in aquaria in order to monitor biological processes and investigate coral behaviour and physiological responses under different experimental treatments. The first challenging aspect is the collection process, which implies the use of oceanographic research vessels in most occasions since these organisms inhabit areas between ca. 150 m to more than 1000 m depth, and specific sampling gears. The next challenge is the maintenance of the animals on board (in situations where cruises may take weeks) and their transport to home laboratories. Maintenance in the home laboratories is also extremely challenging since special conditions and set-ups are needed to conduct experimental studies to obtain information on the biological processes of these animals. The complexity of the natural environment from which the corals were collected cannot be exactly replicated within the laboratory setting; a fact which has led some researchers to question the validity of work and conclusions drawn from such undertakings. It is evident that aquaria experiments cannot perfectly reflect the real environmental and trophic conditions where these organisms occur, but: (1) in most cases we do not have the possibility to obtain equivalent in situ information and (2) even with limitations, they produce relevant information about the biological limits of the species, which is especially valuable when considering potential future climate change scenarios. This chapter includes many contributions from different authors and is envisioned as both to be a practical “handbook” for conducting cold-water coral aquaria work, whilst at the same time offering an overview on the cold-water coral research conducted in Mediterranean laboratories equipped with aquaria infrastructure. Experiences from Atlantic and Pacific laboratories with extensive experience with cold-water coral work have also contributed to this chapter, as their procedures are valuable to any researcher interested in conducting experimental work with cold-water corals in aquaria. It was impossible to include contributions from all laboratories in the world currently working experimentally with cold-water corals in the laboratory, but at the conclusion of the chapter we attempt, to our best of our knowledge, to supply a list of several laboratories with operational cold-water coral aquaria facilities.
    Repository Name: EPIC Alfred Wegener Institut
    Type: Inbook , peerRev
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