ABSTRACT
Rootlets induced from the petiole base of L. purpureus, using IAA and kinetin was used for enhanced multiplication of arbuscular mycorrhizal (AM) fungus, G. deserticula. Using conserved short arbitrary oligonucleotides, as specific primers, we amplified the ITS-region, a molecular marker for fungal identification, from the genomic DNA extracted from cultured spores of G. deserticola, and genomic DNA extracted from the mycelium of L. fraterna. The capacity of fungal colonization and subsequent spore formation of G. deserticola, compared with the natural root system was evaluated. This technology would provide a simple way to multiply AM fungi and to produce spores without microbial contamination useful for further molecular characterization.
Subject(s)
Basidiomycota/genetics , DNA/chemistry , DNA Primers/chemistry , DNA, Fungal/metabolism , Indoleacetic Acids/pharmacology , Microbiology , Mycorrhizae/genetics , Naphthaleneacetic Acids/pharmacology , Oligonucleotides/chemistry , Plant Growth Regulators/pharmacology , Plant Roots/metabolism , Polymerase Chain Reaction , Spores, Fungal/metabolism , Temperature , Time FactorsABSTRACT
High mobility group-1 (HMGB-1) enhances the DNA interactions and possesses a transcriptional activation potential for several families of sequence-specific transcriptional activators. In order to examine the effect of HMGB-1 on the cell cycle progression in MCF-7 cells, the HMGB-1 expression vector was transfected into synchronized MCF-7 cells, and the effect of HMGB-1 overexpression on the cell cycle was examined. The HMGB-1 protein level in the transfected cells increased 4.87-fold compared to the non-transfected cells. There were few changes in the cell cycle phase distribution after HMGB-1 overexpression in the MCF-7 cells. Following the estrogen treatment, the cell cycle progressed in both the HMGB-1 overexpressed MCF-7 and the mock-treated cells. However, a larger proportion of HMGB-1 overexpressing MCF-7 cells progressed to the either S or G2 phase than the mock-treated cells. The mRNA levels of the cell cycle regulators changed after being treated with estrogen in both the HMGB-1 overexpressing MCF-7 and the mock-treated cells, but the changes in the expression level of the cell cycle regulator genes were more prominent in the HMGB-1 overexpressing MCF-7 cells than in the mock-treated cells. In conclusion, HMGB-1 overexpression itself does not alter the MCF-7 cell cycle progression, but the addition of estrogen to the HMGB-1 overexpressing MCF-7 cells appears to accelerate the cell cycle progression.
Subject(s)
Humans , Blotting, Western , Cell Cycle , Cell Line, Tumor , Densitometry , Estrogens/metabolism , G2 Phase , Genetic Vectors , HMGB1 Protein/biosynthesis , Kinetics , Oligonucleotides/chemistry , Plasmids/metabolism , Protein Structure, Tertiary , RNA, Messenger/metabolism , Reverse Transcriptase Polymerase Chain Reaction , S Phase , Time Factors , Transcriptional Activation , TransfectionABSTRACT
A diferencia del resto de las moléculas biológicas, los fosfolípidos son capaces de autoensamblarse espontáneamente. Con ellos es relativamente simples generar estructuras selladas extremadamente estables, de tamaño, forma y empaquetamiento controlables, llamadas liposomas. En este artículo revisaremos el uso de liposomas para generar vectores que mejoren los procesos de transfección en células eucarioticas, tanto in vivo como in vitro. Empleando vectores lipídicos, es potencialmente posible enviar selectivamente un segmento de AND a cualquier sitio del cuerpo, forzarlo a ingresar al interior celular y aun controlar el destino intracelular de la carga transportada. La clave del éxito de la transfección por medio de vectores lipídicos radica en que protegen mecánicamente al AND de la degradación plasmática, ofreciendo a la vez la oportunidad de controlar su biodistribuición, independientemente del tamaño del segmento de AND que se quiera expresar. Asimismo, son no carcinogénicos y pobremente inmunogénicos. Los avances en la química de sintésis de lípidos permitirán construir vectores cada vez más eficientes, que capitan con los altos niveles de transfección de los vectores virales, sumado a las ventajas de extrema versatilidad, facilidad de preparación y bioseguridad propias de la moléculas autoensamblables.
Subject(s)
Animals , Humans , Genetic Therapy , Genetic Vectors , Lipids , Liposomes , Transfection , DNA/chemistry , Lipids/chemistry , Liposomes/chemistry , Nucleic Acids/chemistry , Oligonucleotides/chemistryABSTRACT
The expression of a native form of the foot-and-mouth disease virus RNA polymerase was obtained. Two oligonucleotides of 66 base pairs were used to renuild the 5' end of the gene and to introduce the first methionine codon. The expression of the active polymerase in E. coli was achieved by inserting the gene before the tac promoter of the pKK223-3 plasmid