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991.
Summary A versatile plasmid marker rescue transformation system was developed for homology-facilitated cloning in Bacillus subtilis. It is based on the highly efficient host-vector system 6GM15-pHPS9, which allows the direct selection of recombinants by means of -galactosidase -complementation. The system offers several advantages over previously described cloning systems: (1) the convenient direct selection of recombinants; (2) the ability to effectively transform B. subtilis competent cells with plasmid monomers, which allows the forced cloning of DNA fragments with high efficiency; (3) the availability of 6 unique target sites, which can be used for direct clone selection, SphI, NdeI, NheI, BamHI, SmaI and EcoRI; and (4) the rapid segregational loss of the helper plasmid from the transformed cells. 相似文献
992.
Liposome-mediated gene delivery into plant cells 总被引:1,自引:0,他引:1
Liposomes may offer several advantages as vectors for gene delivery into plant cells: (1) enhanced delivery of encapsulated DNA by membrane fusion, (2) protection of nucleic acids from nuclease activity, (3) targeting to specific cells, (4) delivery into a variety of cell types besides protoplasts by entry through plasmodesmata, (5) delivery of intact small organelles. Realization of these advantages calls for the construction of efficient liposomes, for appropriate fusion conditions and for an understanding of the nature of liposome-cell interactions. Various characteristics and techniques of the liposome-cell system are described (mode of delivery, liposome types and composition, and means of promoting delivery of liposome contents). Data of liposome-mediated delivery of various macromolecules into plant cells, with special reference to protoplasts, calli and pollen are reviewed. This includes data obtained by the use of fluorescent probes, radioactive-labelled DNA, viral nucleic acids and expression of plasmid-DNA. Structure and characteristics of plant surfaces and plasmodesmata are discussed with respect to DNA entry. It is suggested that liposome-mediated gene delivery into plant cells, and not only protoplasts, will be advantageous in certain specific tissues and situations. 相似文献
993.
A transformation and regeneration system has been developed for Nicotiana alata, a plant which is being intensively studied as a model of gametophytic self-incompatibility. Plantlets can be regenerated efficiently from seedling hypocotyls. Kanamycin-resistant, transformed plants have been obtained by cocultivation of regenerating hypocotyls with Agrobacterium tumefaciens strain LBA4404 containing a binary vector. The transformation frequency was low with <1% of tissue explants regenerating transformed plants. The transformed plants contained from one to three copies of the introduced DNA. In most cases, the kanamycin resistance phenotype was transmitted to the offspring as a normal Mendelian factor. In one unusual case, none of the offspring inherited the kanamycin resistance of the transformed maternal parent. This plant may have been chimeric or the kanamycin resistance gene may have been inactivated. 相似文献
994.
Summary Uniformly14C labelled glucose, cellulose and wheat straw and specifically14C labelled lignin component in corn stalks were aerobically incubated for 12 weeks in a chernozem soil alongwith15N labelled ammonium sulphate. Glucose was most readily decomposed, followed in order by cellulose, wheat straw and corn stalk lignins labelled at methoxyl-, side chain 2-and ring-C. More than 50% of14C applied as glucose, cellulose and wheat straw evolved as CO2 during the first week. Lignin however, decomposed relatively slowly. A higher proportion of14C was transformed into microbial biomass whereas lignins contributed a little to this fraction.After 12 weeks of incubation nearly 60% of the lignin14C was found in humic compounds of which more than 70% was resistant to hydrolysis with 6N HCl. Maximum incorporation of15N in humic compounds was observed in cellulose amended soil. However, in this case more than 80% of the15N was in hydrolysable forms.Immobilization-remineralization of applied15N was most rapid in glucose treated soil and a complete immobilization followed by remineralization was observed after 3 days. The process was much slow in soil treated with cellulose, wheat straw or corn stalks. More than 70% of the newly immobilized N was in hydrolysable forms mainly reepresenting the microbial component.Serial hydrolysis of soil at different incubation intervals showed a greater proportion of 6N HCl hydrolysable14C and15N in fractions representing microbial material.14C from lignin carbons was relatively more uniformly distributed in different fractions as compared to glucose, cellulose and wheat straw where a major portion of14C was in easily hydrolysable fractions. 相似文献
995.
The presence of a newly formed primary cell wall was shown to be required for attachment and subsequent transformation of tobacco leaf protoplasts by Agrobacterium tumefaciens in cocultivation experiments. In these experiments both protoplasts at different stages after their isolation and cell-wall inhibitors were used. The specificity of Agrobacterium attachment was shown by using other kinds of bacteria that did not attach. By diminishing the concentration of divalent cations using ethylenediaminetetraacetic acid, neither attachment nor transformation was found; however, when more specifically the Ca2+concentration was lowered by ethylene glycol-bis (-aminoethyl ether)-N,N,N,N-tetraacetic acid, both phenomena occurred. Commercial lectins had no effect on binding, but this observation does not exclude the involvement of other lectins. Protoplasts isolated from various crown-gall callus tissues also developed binding sites, but when they were at the stage of dividing cells, attachment of agrobacteria was no longer observed. In this respect, cells from protoplasts of normal tobacco leaves behaved differently. Even 16 d after protoplast isolation, the dividing cells were still able to bind A. tumefaciens, while transformation was not detected. For transformation of 3-d-old tobacco protoplasts, a minimal co-cultivation period of 24 h was required, while optimal attachment took place within 5 h. It is concluded that the primary cell wall was sufficiently well formed that certain functional receptor molecules were available for attachment of Agrobacterium as the first step of a multistep process leading to the transformation of cells. The expression of bacterial functions required for attachment, moreover, was independent of the presence of Ti-plasmid.Abbreviations ConA
concanavalin A
- CW
calcofluor white
- EDTA
ethylenediaminetetraacetic acid
- EGTA
ethylene glycol-bis (-aminoethyl ether)-N,N,N,N-tetraacetic acid
- -Man
-methyl-d-mannoside 相似文献
996.
The ultrastructural and biochemicalphysiological aspects of postfloral greening have been studied in hypsophylls of Heliconia aurantiaca Ghiesbr., Guzmania cf. x magnifica Richter and Spathiphyllum wallisii Regel. In all three species the greening of the hypsophylls is due to plastid transformation, chloroplast formation proceeding from the initially different types of plastids. The degradation process of the original plastid structures and the mode of thylakoid formation are distinct in each case. In none of the species do the transformed plastids look identical to the chloroplasts of the corresponding foliage leaves. On a chlorophyll basis, the rate of photosynthesis of the greened hypsophylls surpasses the rate of the leaves considerably in Spathiphyllum, but is much lower in Heliconia (no data for Guzmania). In all species, anatomy, plastid structure, pigments, 77° K-fluorescence emission, ribulose-1,5-bis-phosphate carboxylase activities and short-term photosynthesis 14CO2-assimilation patterns prove the greened hypsophylls to be capable of providing additional carbon to the developing fruits, thus supplementing the import of organic matter from the foliage leaves.Abbreviations MDH
malate dehydrogenase (EC 1.1.1.37)
- PEPCase
phosphoenolpyruvate carboxylase (EC 4.1.1.31)
- RuBPCase
ribulose-1,5-bisphosphate carboxylase (EC 4.1.1.39) 相似文献
997.
Summary Mutations of the homeotic gene fork head (fkh) of Drosophila transform the non-segmented terminal regions of the embryonic ectoderm into segmental derivatives: Pre-oral head structures and the foregut are replaced by post-oral head structures which are occasionally associated with thoracic structures. Posterior tail structures including the hindgut and the Malpighian tubules are replaced by post-oral head structures associated with anterior tail structures. The fkh gene shows no maternal effect and is required only during embryogenesis. The phenotypes of double mutants indicate that fkh acts independently of other homeotic genes (ANT-C, BX-C, spalt) and caudal. In addition, the fkh domains are not expanded in Polycomb (Pc) group mutant embryos. Ectopic expression of the homeotic selector genes of the ANT-C and BX-C in Pc group mutant embryos causes segmental transformations in terminal regions of the embryo only in the absence of fkh gene activity. Thus, fkh is a region-specific homeotic rather than a selector gene, which promotes terminal as opposed to segmental development.
Offprint requests to: Institut für Biologie II (Genetik), Universität Tübingen, Auf der Morgenstelle 28, D-7400 Tübingen, Federal Republic of Germany 相似文献
998.
999.
Transformation of plants via the shoot apex 总被引:4,自引:0,他引:4
E. C. Ulian R. H. Smith J. H. Gould T. D. McKnight 《In vitro cellular & developmental biology. Plant》1988,24(9):951-954
Summary We have transformed petunia byAgrobacterium tumefaciens containing genes for kanamycin resistance and beta-glucuronidase using isolated shoot apices from seedling tissue. Regeneration
of transformed plants in this model system was rapid. The technique of shoot apex transformation is an alternative for use
inAgrobacterium-mediated transformation of dicotyledonous crop species for which a method of regeneration via protoplasts, leaf disks, or
epidermal strips does not exist. This approach offers direct and rapid regeneration of plants and low risk of tissue-culture-induced
genetic variation.
Texas Agricultural Experiment Station Technical Article No. 23317. 相似文献
1000.
Joseph R. Perera Alexander V. Glasunov Vadim M. Glaser Alla V. Boreiko 《Molecular & general genetics : MGG》1988,213(2-3):421-424
Summary We studied the repair of double-strand breaks (DSB) in plasmid DNA introduced into haploid cells of the yeast Saccharomyces cerevisiae. The efficiency of repair was estimated from the frequency of transformation of the cells by an autonomously replicated linearized plasmid. The frequency of lithium transformation of Rad+ cells was increased greatly (by 1 order of magnitude and more) compared with that for circular DNA if the plasmid was initially linearized at the XhoI site within the LYS2 gene. This effect is due to recombinational repair of the plasmid DNA. Mutations rad52, rad53, rad54 and rad57 suppress the repair of DSB in plasmid DNA. The kinetics of DSB repair in plasmid DNA are biphasic: the first phase is completed within 1 h and the second within 14–18 h of incubating cells on selective medium. 相似文献