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Genetic Engineering Publications - GEG Tech top picks
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BigField GEG Tech's insight:
Design of new tools for genome editing. http://geg-tech.com/products/vectors.html?applications=Recombinase%2C+editors 
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PhiC31 Integrase Induces Efficient Site-Specific Recombination in the Capra hircus Genome | Abstract
BigField GEG Tech's insight:
Proof of concept of how PhiC31 recombinase works in Capra hircus fibroblasts. Moreover, this study points out new pseudo attP sites in the mammalian genome.
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PhiC31 integrase-mediated gene delivery has been extensively used in gene therapy and animal transgenesis. However, random integration events are observed in phiC31-mediated integration in different types of mammalian cells; as a result, the efficiencies of pseudo attP site integration and evaluation of site-specific integration are compromised. To improve this system, we used an attB-TK fusion gene as a negative selection marker, thereby eliminating random integration during phiC31-mediated transfection. We also excised the selection system and plasmid bacterial backbone by using two other site-specific recombinases, Cre and Dre. Thus, we generated clean transgenic bovine fetal fibroblast cells free of selectable marker and plasmid bacterial backbone. These clean cells were used as donor nuclei for somatic cell nuclear transfer (SCNT), indicating a similar developmental competence of SCNT embryos to that of non-transgenic cells. Therefore, the present gene delivery system facilitated the development of gene therapy and agricultural biotechnology.
BigField GEG Tech's insight:
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BigField GEG Tech's insight:
The authors used phiC31 integrase to insert a copy of a β-globin / puromycin (purr) vector into specific genomic locations of a human hematopoietic cell line. After genomic integration, they used Cre recombinase to remove the bacterial backbone and purr.Following that, the stable β-chain expression was continued for several months in the absence of selective pressure.
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BigField GEG Tech's insight:
The authors enhanced the integration site-specificity of the phiC31 integrase-based vector using a sequence-specific DNA-binding protein containing Gal4 and LexA DNA-binding motifs. In human cells, they showed thanks to quantitative PCR that integration around the LexA operator was 26-fold higher than with the control.
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PLOS ONE: an inclusive, peer-reviewed, open-access resource from the PUBLIC LIBRARY OF SCIENCE. Reports of well-performed scientific studies from all disciplines freely available to the whole world.
BigField GEG Tech's insight:
Genetic engineering of iPs invloving 3 site specific recombinases. PhiC31 integrase was used to mediate initial placement of a single copy of a reprogramming plasmid into the genome at a safe location. A second phage integrase, Bxb1, was used to place the full-length dystrophin coding sequence into the same location, and Cre resolvase was utilized to excise unwanted sequences.
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![Long-term and efficient expression of human [beta]-globin gene in a hematopoietic cell line using a new site-specific integrating non-viral system | Genetic Engineering Publications - GEG Tech top picks | Scoop.it](https://img.scoop.it/McIgc7_5fF4eSerUlFxk4jl72eJkfbmt4t8yenImKBVvK0kTmF0xjctABnaLJIm9)
In this work, the scientists have developed a novel system to convert scFvs from a phage display vector directly into IgGs without any in vitro subcloning steps. This new vector system, named pMINERVA, makes clever use of site-specific bacteriophage integrases that are expressed in Escherichia coli and intron splicing that occurs within mammalian cells.