Cell type differences in activity of the Streptomyces bacteriophage phiC31 integrase.

Nucleic Acids Research
Christof MauckschCarsten Rudolph

Abstract

Genomic integration by the Streptomyces bacteriophage C31 integrase is a promising tool for non-viral gene therapy of various genetic disorders. We investigated the C31 integrase recombination activity in T cell derived cell lines, primary T lymphocytes and CD34(+) haematopoietic stem cells in comparison to mesenchymal stem cells and cell lines derived from lung-, liver- and cervix-tissue. In T cell lines, enhanced long-term expression above control was observed only with high amounts of integrase mRNA. Transfections of C31 integrase plasmids were not capable of mediating enhanced long-term transgene expression in T cell lines. In contrast, moderate to high efficiency could be detected in human mesenchymal stem cells, human lung, liver and cervix carcinoma cell lines. Up to 100-fold higher levels of recombination product was found in C31 integrase transfected A549 lung than Jurkat T cells. When the C31 integrase activity was normalized to the intracellular integrase mRNA levels, a 16-fold difference was found. As one possible inhibitor of the C31 integrase, we found 3- to 5-fold higher DAXX levels in Jurkat than in A549 cells, which could in addition to other yet unknown factors explain the observed discrepancy of C31 integrase...Continue Reading

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Citations

Mar 16, 2012·Applied Microbiology and Biotechnology·Yuan-Yuan XueJin-Zhong Chen
Oct 17, 2009·Molecular Genetics and Genomics : MGG·Kentaro MoritaHideo Takahashi
Dec 1, 2012·Human Gene Therapy Methods·Marc-André RobertRénald Gilbert
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Jul 25, 2009·FEMS Microbiology Letters·Kentaro MoritaHideo Takahashi

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Methods Mentioned

BETA
transfection
density gradient centrifugation
transfections
protein assay
electrophoresis
PCR
transfect

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