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1

Effectiveness of Agrobacterium-mediated transformation in selected forms of tomato (Lycopersicon esculentum Mill.) and preliminary characteristics of transgenic plants.

100%
Bartoszewski G., Malepszy S., Niemirowicz-Szczytt K.
|
EN
Three forms of Lycopersicon esculentum Mill. variety Beta, line ls and line nor were subject to Agrobacterium-mediated transformation with two constructs: pBI121 and pRUR528. In both cases transgenic plants were obtained which formed roots in a medium with kanamycin and showed expected band in the PCR analysis. The effectiveness of the transformation, measured as the number of explants forming structures on selection medium, was different for the used genotypes when a bacterial strain carrying plasmid pRUR528 was used. No genotype-dependent difference in effectiveness of the transformation was observed in the case of pBI121. The genotype-dependent differences in the effectiveness of the transgenic shoots regeneration and rooting were observed. 76% of transgenic plants showed unchanged ploidy level.
2

Agrobacterium-mediated transformation of plants - basic model of the process and key factors involved in it

100%
Nadolska-Orczyk A.
Biotechnologia
|
1999
|
issue 1
116-124
EN
During infection of plant tissue by Agrobacterium, a fragment of Ti plasmid (T-DNA) is transferred to a cell. T-DNA is delimited by 25 bp direct repeats, which are necessary to direct it. Any piece of DNA flanked by these sequences can be transferred to the plant cell, where it becomes integrated into the plant genome. The processing and transfer of T-DNA are mediated by vir region localized on the Ti-plasmid. Based on the summarized model of plant transformation via Agrobacterium, the key factors involved in this mechanism are reviewed. There are two basic components of the process: Agro strain with its vector system and recipient, usually totipotent plant cell. The most important factors, which should be taken into consideration, include: Agrobacterium host specificity and its vector system, tissue culture techniques and their limitations, chemical agents inducing virulence genes and selection strategies. Some of them can influence the expression pattern of the introduced genes.
3

Agrobacterium-mediated tomato transformation

80%
Bartoszewski G., Niemirowicz-Szczytt K.
Biotechnologia
|
1998
|
issue 1
43-63
EN
Tomato is one of the model species used for crop transformation. The first transgenic tomato variety, known as Flavr Savr?, was approved for sale in the USA in 1994. The introduced trait of this cultivar is delayed ripening. In 1996, its acreage was reported to be 10,000 acres. Another variety characterised by delayed ripening is Endless Summer?, approved in 1995. There are some other cultivars with new traits, such as thicker skin, altered pectin or resistance to viruses (TSWV, ToMV) either approved or pending approval. In addition, a wide range of basic research on tomato transformation has been carried out, including studies on resistance to herbicides, viruses, fungi, bacteria, insects as well as on altered transcription regulation, ripening, carotenoid synthesis, level of auxines and cytokinines, carbohydrates, proteins and specific vaccines. Further improvement of tomato varieties is expected with the use of Agrobacterium tumefaciens.
4

Cucumber transformation methods - the review

80%
Yin Z., Bartoszewski G., Szwacka M., Malepszy S.
Biotechnologia
|
2005
|
issue 1
95-113
EN
Several aspects of cucumber transformation including the ways of transgene introduction, factors influencing the transformation efficiency and the fate of the introduced genes were reviewed. Various transgenes have been introduced into the cucumber genome mostly via the Agrobacterium-mediated transformation. The frequency of Agrobacterium-mediated transformation ranged from 0.8 to 10% and was influenced by the selection agent, the regeneration efficiency, activation of vir genes expression, the explant size, bacteria cell density, the length of exposure and the co-cultivation period. The transgenes were integrated mostly as single copy in the Agrobacterium-mediated transformation and as multiple copies in direct transformation. Variable levels of the transgene expression were observed. The transmission of the transgenes as well as the transgenic phenotype follow the Mendelian, and rarely non-Mendelian, ratio. The production of marker-free transgenic cucumber and use of an alternative transformation method are recommended.
5

Transfer and transgene expression

80%
Szopa J., Wrobel M.
Biotechnologia
|
2001
|
issue 1
63-72
EN
The development of transgenic plants strongly depends on the stable introduction of foreign nucleic acid into the plant genome. Recently, various transformation methods have been developed and successfully used for the development of transgenic plants of agricultural importance. Various vectors have been prepared, however the binary vector based on agrobacterial T-DNA is most commonly used. Of vectorless gene transfer systems, particle bombardment seems to be most useful. In both approaches, stable integration of DNA is based on random hybridization. The transformation efficiency is measured by the level of transgene expression and it may be potentially improved by different modifications, including insertion of multiple copies of promoter into a particular gene, insertion of procaryotic enhancer, increasing of mRNA stability, insertion of SAR/MAR sequences at the ends of gene, etc. Very recently it has been found that the increase of histone synthesis enhances transgene expression possibly as a result of increased transformation efficiency. Transgenic plants production also depends on the efficiency of the plant regeneration system which is used. There is no universally applicable method for regeneration of different tissues from various sources, thus regeneration protocol should be modified appropriately for each tissue and species.
6

Transformation of selected genotypes of tomato with the help of A. tumefaciens

80%
Bartoszewski G., Fedorowicz O., Malinowski R., Niedziela A., Niemirowicz-Szczytt K.
Biotechnologia
|
2000
|
issue 4
99-102
EN
Tomato belongs to important crops widely cultivated all over the world. It is also one of the five most popular vegetables grown in Poland. At the same time, tomato is known to be a model species in modern biology and biotechnology. Since 1985 a lot of reports on tomato transformation with the use of Agrobacterium have been published. Recently, first transgenic varieties of this species have also been developed. Flavr Savr? obtained by Calgene, USA was the first cultivar obtained as a result of genetic engineering, officially registered in the United States. In our Department the methods of tomato transformation with Agrobacterium tumefaciens have been adapted and optimised. Numerous transgenic plants have been obtained, such as commercial varieties (Beta, Potentat), inbred lines as well as tomato mutants (non-ripening, lateral suppressor). The following genes were introduced to the above forms: beta-glucuronidase reporter gene (gusA), thaumatin gene (sweet protein), isopentenyl transferase gene (ipt, coding the key enzyme in cytokinin metabolic pathway) and thus homozygous lines were developed (T2 generation). Most recently, attempts have been made to incorporate mgfp5-ER gene coding green fluorescence protein, nucleoprotein (N) gene from tomato spotted wilt virus (TSWV) and cDNA of putative P450 cytochrome (CYP72) in order to test gene expression and interaction.
7

Effectivity of plant genetic transformation systems and proteins-nucleic acids interaction

70%
Ciesielczyk P., Bartkowiak S.
Biotechnologia
|
1995
|
issue 2
52-72
EN
Possible interactions between proteins and nucleic acids which might contribute to general effectivity op plant genetic transformation systems are reviwed. Experimental data concerning direct and A tumefaciens - based plant genetic transformation systems are compared with data obtained from research on nucleocytoplasmic exchange machanisms of biopolmers.In thaht context, mechanisms of hnRNA interactions with hnRNP appear to be of great importance. This type of interactions is determined by conservative nucleic acida bindng domains of many cellular proteins e.g. RGG and RBD.These proteins seem to play an important role in the integration of many other cellular processes.
8

Transformation of microspore-derived embryos of winter oilseed rape (Brassica napus L.) by using Agrobacterium tumefaciens

61%
Cegielska-Taras T., Pniewski T., Szala L.
Journal of Applied Genetics
|
2008
|
vol. 49
|
issue 4
343-347
EN
Haploid microspore-derived embryos (MDEs) constitute a unique material for the introduction of new traits into winter oilseed rape (Brassica napus). MDEs have been transformed by using Agrobacterium tumefaciens strains EHA105 and LBA4404, both carrying the binary vector pKGIB containing the uidA gene encoding -glucuronidase (GUS) and the bar gene as a marker of resistance to phosphinotricin. Transformed embryos expressed GUS and regenerated plants that were resistant to herbicide Basta, as confirmed by a leaf-painting test. Progeny plants of the transformant T-39 were all transgenic, as they inherited T-DNA from their doubled haploid parental plant. Southern-blot analysis confirmed the integration and transmission of T-DNA into T1 plants. Transformation of MDEs facilitates the obtaining of winter oilseed rape homozygous for the introduced genes.
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