RESUMO
To produce transgenic plants in current biotechnology selectable marker genes are used that lead to the selectivity.of transformants from non-transformed organisms. However, after the transgenic event has been occurred, the presence of these genes in transformed genome in general is uselless. Moreover, the continued presence of this kind of genes in transgenic plants with their further commercialization may raise certain public concern. Therefore, various techniques have been developed in recent years to obtain marker free transgenic plants. In the present review the main strategies for removal of selective marker DNA sequences that are used in genetic engineering are described. The most popular among them is site-specific recombination technology. The particular attention is paid to site-specific recombinase system Cre/loxP. The using of a new approach with site-specific recombinase system Cre/loxP under the control of 35S promoter to generate marker-free genetically modified plants is described.
Assuntos
Biotecnologia/métodos , DNA Nucleotidiltransferases/genética , Engenharia Genética/métodos , Plantas Geneticamente Modificadas/enzimologia , Plantas Geneticamente Modificadas/genética , Transformação Genética , Marcadores Genéticos , Regiões Promotoras GenéticasRESUMO
The protocol for production of full-size cDNA using SuperScript Full-Length cDNA Library Construction Kit II (Invitrogen) was tested and high quality cDNA library from meristematic tissue of finger millet panicle (Eleusine coracana (L.) Gaertn) was created. The titer of obtained cDNA library comprised 3.01 x 10(5) CFU/ml in avarage. In average the length of cDNA insertion consisted about 1070 base pairs, the effectivity of cDNA fragment insertions--99.5%. The selective sequencing of cDNA clones from created library was performed. The sequences of cDNA clones were identified with usage of BLAST-search. The results of cDNA library analysis and selective sequencing represents prove good functionality and full length character of inserted cDNA clones. Obtained cDNA library from meristematic tissue of finger millet panicle represents good and valuable source for isolation and identification of key genes regulating metabolism and meristematic development and for mining of new molecular markers to conduct out high quality genetic investigations and molecular breeding as well.
Assuntos
DNA Complementar/genética , DNA de Plantas/genética , Eleusine/genética , Biblioteca Gênica , Meristema/genética , Sequência de Bases , Clonagem Molecular , Primers do DNA/genética , DNA de Plantas/isolamento & purificação , Escherichia coli/genética , Dados de Sequência Molecular , RNA de Plantas/genética , RNA de Plantas/isolamento & purificaçãoRESUMO
Comparative study of long-term existing genome damages in the chronically irradiated pine seeds from different parts of Chernobyl zone have been performed by a pulsed field gel electrophoresis and hydroxyapatite chromatography. Some differences were established for DNA degradation kinetic from protoplasts with various accumulated doses of irradiation. Results obtained could testify the existence of long-living changes in the experimental plants genome.
