RNA as a source of transposase for Sleeping Beauty-mediated gene insertion and expression in somatic cells and tissues

Andrew Wilber; Joel L. Frandsen; Jennifer L. Geurts; David A. Largaespada; Perry B. Hackett; R. Scott McIvor

doi:10.1016/j.ymthe.2005.10.014

RNA as a source of transposase for Sleeping Beauty-mediated gene insertion and expression in somatic cells and tissues

Andrew Wilber
, Joel L. Frandsen
, Jennifer L. Geurts
, David A. Largaespada
, Perry B. Hackett
, R. Scott McIvor

Research output: Contribution to journal › Article › peer-review

109 Scopus citations

Abstract

Sleeping Beauty (SB) is a DNA transposon capable of mediating gene insertion and long-term expression in vertebrate cells when co-delivered with a source of transposase. In all previous reports of SB-mediated gene insertion in somatic cells, the transposase component has been provided by expression of a co-delivered DNA molecule that has the potential for integration into the host cell genome. Integration and continued expression of a gene encoding SB transposase could be problematic if it led to transposon re-mobilization and reintegration. We addressed this potential problem by supplying the transposase-encoding molecule in the form of mRNA. We show that transposase-encoding mRNA can effectively mediate transposition in vitro in HT1080 cells and in vivo in mouse liver following co-delivery with a recoverable transposon or with a luciferase transposon. We conclude that in vitro-transcribed mRNA can be used as an effective source of transposase for SB-mediated transposition in mammalian cells and tissues.

Original language	English (US)
Pages (from-to)	625-630
Number of pages	6
Journal	Molecular Therapy
Volume	13
Issue number	3
DOIs	https://doi.org/10.1016/j.ymthe.2005.10.014
State	Published - Mar 2006

Bibliographical note

Funding Information:
We thank Aron Geurts, Corey Carlson, and the Biological Imaging and Processing Laboratory at the University of Minnesota for technical assistance. This work was supported by a grant from the Arnold and Mabel Beckman Foundation (R.S.M) and Training Grant T32 GM08347 from NIGMS (A.W.). A.W. is the recipient of a University of Minnesota Doctoral Dissertation Fellowship.

Keywords

Bioluminescent imaging
Liver
Non-viral integration
RNA delivery
Sleeping Beauty

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10.1016/j.ymthe.2005.10.014

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title = "RNA as a source of transposase for Sleeping Beauty-mediated gene insertion and expression in somatic cells and tissues",

abstract = "Sleeping Beauty (SB) is a DNA transposon capable of mediating gene insertion and long-term expression in vertebrate cells when co-delivered with a source of transposase. In all previous reports of SB-mediated gene insertion in somatic cells, the transposase component has been provided by expression of a co-delivered DNA molecule that has the potential for integration into the host cell genome. Integration and continued expression of a gene encoding SB transposase could be problematic if it led to transposon re-mobilization and reintegration. We addressed this potential problem by supplying the transposase-encoding molecule in the form of mRNA. We show that transposase-encoding mRNA can effectively mediate transposition in vitro in HT1080 cells and in vivo in mouse liver following co-delivery with a recoverable transposon or with a luciferase transposon. We conclude that in vitro-transcribed mRNA can be used as an effective source of transposase for SB-mediated transposition in mammalian cells and tissues.",

keywords = "Bioluminescent imaging, Liver, Non-viral integration, RNA delivery, Sleeping Beauty",

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note = "Funding Information: We thank Aron Geurts, Corey Carlson, and the Biological Imaging and Processing Laboratory at the University of Minnesota for technical assistance. This work was supported by a grant from the Arnold and Mabel Beckman Foundation (R.S.M) and Training Grant T32 GM08347 from NIGMS (A.W.). A.W. is the recipient of a University of Minnesota Doctoral Dissertation Fellowship.",

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AU - Frandsen, Joel L.

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AU - Largaespada, David A.

AU - Hackett, Perry B.

AU - McIvor, R. Scott

N1 - Funding Information: We thank Aron Geurts, Corey Carlson, and the Biological Imaging and Processing Laboratory at the University of Minnesota for technical assistance. This work was supported by a grant from the Arnold and Mabel Beckman Foundation (R.S.M) and Training Grant T32 GM08347 from NIGMS (A.W.). A.W. is the recipient of a University of Minnesota Doctoral Dissertation Fellowship.

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N2 - Sleeping Beauty (SB) is a DNA transposon capable of mediating gene insertion and long-term expression in vertebrate cells when co-delivered with a source of transposase. In all previous reports of SB-mediated gene insertion in somatic cells, the transposase component has been provided by expression of a co-delivered DNA molecule that has the potential for integration into the host cell genome. Integration and continued expression of a gene encoding SB transposase could be problematic if it led to transposon re-mobilization and reintegration. We addressed this potential problem by supplying the transposase-encoding molecule in the form of mRNA. We show that transposase-encoding mRNA can effectively mediate transposition in vitro in HT1080 cells and in vivo in mouse liver following co-delivery with a recoverable transposon or with a luciferase transposon. We conclude that in vitro-transcribed mRNA can be used as an effective source of transposase for SB-mediated transposition in mammalian cells and tissues.

AB - Sleeping Beauty (SB) is a DNA transposon capable of mediating gene insertion and long-term expression in vertebrate cells when co-delivered with a source of transposase. In all previous reports of SB-mediated gene insertion in somatic cells, the transposase component has been provided by expression of a co-delivered DNA molecule that has the potential for integration into the host cell genome. Integration and continued expression of a gene encoding SB transposase could be problematic if it led to transposon re-mobilization and reintegration. We addressed this potential problem by supplying the transposase-encoding molecule in the form of mRNA. We show that transposase-encoding mRNA can effectively mediate transposition in vitro in HT1080 cells and in vivo in mouse liver following co-delivery with a recoverable transposon or with a luciferase transposon. We conclude that in vitro-transcribed mRNA can be used as an effective source of transposase for SB-mediated transposition in mammalian cells and tissues.

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KW - Liver

KW - Non-viral integration

KW - RNA delivery

KW - Sleeping Beauty

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RNA as a source of transposase for Sleeping Beauty-mediated gene insertion and expression in somatic cells and tissues

Abstract

Bibliographical note

Keywords

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