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EN
Matrix attachment regions (MARs) are thought to participate in the organization and segregation of independent chromosomal loop domains. Although there are several reports on the action of natural MARs in the context of heterologous genes in transgenic plants, in our study we tested a synthetic MAR (sMAR) with the special property of unpairing when under superhelical strain, for its effect on reporter gene expression in tobacco plants. The synthetic MAR was a multimer of a short sequence from the MAR 3' end of the immunoglobulin heavy chain (IgH) enhancer. This sMAR sequence was used to flank the β-glucuronidase (GUS) reporter gene within the T-DNA of the binary vector pBI121. Vectors with or without the sMARs were then used to transform tobacco plants by Agrobacterium tumefaciens. Transgenic plants containing the sMAR sequences flanking the GUS gene exhibited higher levels of transgene expression compared with transgenic plants which lacked the sMARs. This effect was observed independently of the position of the sMAR at the 5' side of the reporter gene. However, variation of the detected transgene expression was significant in all transformed plant populations, irrespective of the construct used. Most genes whose expression has been studied in transgenic plants are generally expressed in appropriate patterns. However, transgene expression can vary within an extremely wide range, often showing only a very low level [1, 2]. Variation in transgene expression is frequently attributed to corresponding variation in the transcription potential of different chromosomal insertion sites. DNA sequences called scaffold/matrix attachment regions (S/MARs) are involved in the structural and functional organization of all eukaryotic genomes. Evolutionarily, the structures of these sequences seem to be conserved. Typically, S/MARs are located every 5 to 200 kb of sequence and are known to bind specifically to components of the nuclear scaffold, therefore suggesting that they are responsible for loop domain base formation [3, 4].Of the MAR elements reported, many do not display extensive sequence homology. It is therefore reasonable to assume that the scaffold probably recognizes some structural features of the MAR DNA rather than a specific sequence [5].MARs appear to be functionally conserved, since animal MARs can bind to plant nuclear scaffolds and vice versa [6, 7].Most MARs have been generally characterized as AT-rich sequences. However, AT-richness per se is not a sufficient criterion for specific sequence recognition of MARs by specific binding proteins [7]. Their capacity to bind to the nuclear matrix is determined by the specific structure of DNA. A prominent structural characteristic of different MARs is their strong potential for extensive unpairing when subjected to superhelical strain [8, 9]. The ability to assume a stably unpaired conformation has been described for several MARs. For example, within the MAR 3' end of immunoglobulin heavy chain (IgH) enhancer there is an AATATATTT motif that is a nucleation site for DNA unwinding [10]. Concatamerized oligonucleotides containing seven repeats of this sequence exhibited a strong affinity for the nuclear scaffold and increased SV40 promoter activity in stably transformed mouse cells [11].In this paper we present results of our studies that concern the effect of a synthetic MAR on transgene expression in tobacco plants. The synthetic MAR sequences were used to flank the β-glucuronidase (GUS) gene whose transcription was under control of the 35S CaMV promoter in the binary vector pBI121. This construct was introduced into tobacco plants and the GUS reporter gene expression was monitored in stably transformed plants.
EN
Plant expression vector pBI 121 containing the gene encoding coat protein of Plum Pox Virus of the Skierniewice isolate (CP PPV-S) was prepared (clone pCMl). The construct was used for transformation of Nicotiana tabacum plants using an Agrobacterium tumefaciens based system. About 82% of kanamycin resistant plant lines contained a transgene (the sequence of CP PPV-S) but only 81% of them actively expressed the PPV-S coat protein gene as measured by RT-PCR.
EN
Several methods of transformation are currently available for delivering exogenous DNA to plant cells. Agrobacterium-mediated transformation, microprojectile bombardment and direct protoplast transformation are routinely used today. However, each of them has certain disadvantages, which led to research into the development of novel alternative systems such as infiltration, electroporation of cells and tissues, electrophoresis of embryos, microinjection, pollen-tube pathway, silicon carbide- and liposome-mediated transformation. The low efficiency of transformation is considered to be the main reason for the limited popularity of the alternative transformation methods, other than infiltration and silicon carbide-mediated transformation, which seem to be the most promising ones for practice.
EN
Matrix attachment regions (MARs) are thought to participate in the organization and segregation of independent chromosomal loop domains. Although there are sev­eral reports on the action of natural MARs in the context of heterologous genes in transgenic plants, in our study we tested a synthetic MAR (sMAR) with the special property of unpairing when under superhelical strain, for its effect on reporter gene expression in tobacco plants. The synthetic MAR was a multimer of a short sequence from the MAR 3 end of the immunoglobulin heavy chain (IgH) enhancer. This sMAR sequence was used to flank the β-glucuronidase (GUS) reporter gene within the T-DNA of the binary vector pBI121. Vectors with or without the sMARs were then used to transform tobacco plants by Agrobacterium tumefaciens. Transgenic plants containing the sMAR sequences flanking the GUS gene exhibited higher levels of transgene expression compared with transgenic plants which lacked the sMARs. This effect was observed independently of the position of the sMAR at the 5 side of the re­porter gene. However, variation of the detected transgene expression was significant in all transformed plant populations, irrespective of the construct used.
EN
Standard program of plant protection against fire blight consists of use of :management practices and chemical control method. Recently a new, non-conventional possibilities based on application of biocontrol agents (two biopreparations have been already introduced into practice: Bliteban A506 Pseudomonas fluorescens) and BIossomBless (Pantoea agglomerans), plant active against Erwinia amylovora (AkseBio containing extracts from spicata and Biomit Plussz with extracts from various plant species and ents) and resistance inducers (Regalis, Bion and plant extracts) are of great interest. Also plant transformation with resistance genes such as: hrpN (harpin), dpo (EPS depolymerase) and lytic protein genes (attacin E, cecropin 'SB-37, T4 lysozyme) is a promising perspective.
PL
Standardowy program ochrony roślin przed zarazą ogniową obejmuje stosowanie metody agrotechnicznej i chemicznej. Ostatnio duże zainteresowanie wzbudzają niekonwencjonalne możliwości walki z chorobą obejmujące wykorzystanie czynników biologicznych (opracowano już i wprowadzono do praktyki sadowniczej preparaty Bliteban A506 na bazie bakterii Pseudomonas fluoresceins oraz BlossomBless na bazie Pantoea agglomerans), ekstraktów z roślin zawierających substancje aktywne w stosunku do Erwinia amylovora (m.in. opracowano preparat AkseBio zawierający ekstrakty z Thymbra spicata oraz Biomit Plussz z ekstraktami z różnych roślin i mikroelementami), indukowanie odporności roślin na chorobę (preparaty Regalis i Bion oraz ekstrakty roślin), a także transformację roślin genami kodującymi syntezę harpiny (hrpN), depolimerazę EPS (dpo), białka lityczne (cekropinę, atacynę, lizozym bakteriofaga T4).
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