when a plant is initially transformed, the transgene is present in a heterozygous form. Since white clover is outcrossing and predominantly self incompatible it is necessary to cross primary transgenic plants to produce an BC1 population, then intercross BC1 transgenic plants to produce an F2 population. The F2 population will have a highly heterogeneous genetic background and a variable alleli...

Introduction of DNA sequences into the genome often results in homology-dependent gene silencing in organisms as diverse as plants, fungi, flies, nematodes, and mammals. We previously showed in Cryptococcus neoformans that a repeat transgene array can induce gene silencing at a high frequency during mating (∼50%), but at a much lower frequency during vegetative growth (∼0.2%). Here we report a ...

The bacterial backbone (BB) sequences contained within a canonical plasmid DNA dampen exogenous transgene expression by tenfold to 1,000-fold over a period of a few weeks following transfection into quiescent tissues such as the liver. Minicircle DNA vectors devoid of bacterial plasmid backbone sequences overcome transgene silencing providing persistent transgene expression. Because, we recentl...

High stability of transgene expression is essential for functional genomics studies using transformation approaches and for application of genetic engineering to commercial forestry. We quantified expression of two reporter genes, green fluorescent protein (GFP) and the herbicide bialaphos resistance gene (BAR), in 2256 transgenic poplar trees derived from 404 primary events, and in 106 in vitr...

A high-level expression of a transgene, Ead, encoding the I-Ed alpha-chain is very effective in protection against murine lupus. To investigate the specific contribution of select H-2 haplotypes on the Ead transgene-mediated disease-suppressing effect, we generated H-2 congenic (NZB x BXSB)F1 hybrid mice bearing either H-2b/b, H-2d/b, or H-2d/d haplotype, and compared the transgene-mediated pro...

For safe and effective gene therapy, targeted tissue-restricted transgene expression is desirable. Various methods have been developed to achieve such expression, including the use of tissue-specific promoters. In addition to these approaches, a new system which can regulate transgene expression, including viral gene expression, by exploiting microRNAs (miRNAs) has recently been developed. miRN...

The introduction of foreign genes into early mouse embryos and embryonic stem (ES) cells is invaluable for the analysis of gene function and regulation in the living animal. The use of vectors derived from retroviruses as gene transfer vehicles in this setting has had limited success because of silencing of transgene expression. Here, we show that vectors derived from lentiviruses, which are co...

Helper-dependent adenoviral vectors (HDAd) that express certain transgene products are impossible to produce because the transgene product is toxic to the producer cells, especially when made in large amounts during vector production. Downregulating transgene expression from the HDAd during vector production is a way to solve this problem. In this report, we show that this can be accomplished b...

Somatic cells can be reprogrammed to induced hepatocyte-like cells (iHeps) by overexpressing certain defined factors in direct reprogramming techniques. Of the various methods to deliver genes into cells, typically used genome-integrating viral vectors are associated with integration-related adverse events such as mutagenesis, whereas non-integrating viral vectors have low efficiency, making vi...

Transgenic mice overexpressing approximately eightfold the mouse (Mo) prion protein (PrP) gene carrying the P102L mutation of GSS developed neurodegeneration between 150 and 300 days of age, while controls expressing the wild-type MoPrP-A transgene at the same level remained healthy. Mice overexpressing the wild-type MoPrP-A transgene were highly susceptible to inoculated mouse prions, exhibiti...