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JoVE Journal
Developmental Biology
An Optimized Mouse Embryonic Stem Cell Based Reverse Poly-Transfection Technique for Rapid Explor...
An Optimized Mouse Embryonic Stem Cell Based Reverse Poly-Transfection Technique for Rapid Explor...
JoVE Journal
Developmental Biology
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JoVE Journal Developmental Biology
An Optimized Mouse Embryonic Stem Cell Based Reverse Poly-Transfection Technique for Rapid Exploration of Nucleic Acid Ratios

An Optimized Mouse Embryonic Stem Cell Based Reverse Poly-Transfection Technique for Rapid Exploration of Nucleic Acid Ratios

Full Text
1,594 Views
06:04 min
December 8, 2023

DOI: 10.3791/65766-v

Kieran Maheden1, Karen Hwang1, Ipek Egilmez1, Nika Shakiba1

1School of Biomedical Engineering,University of British Columbia

The present protocol describes a method for reverse poly-transfection of mouse embryonic stem cells during culture with 2i and LIF media. This method yields higher viability and efficiency than traditional forward transfection protocols, while also enabling one-pot optimization of plasmid ratios.

Our work generally focuses on using synthetic genetic devices to manipulate stem cell faith and behavior. By controlling cellular states, we aim to advance the development of next generation cellular therapies. Recently, a deeper understanding of the relationship between the dosage of a faith determining gene and the cells lineage has been established.

This was facilitated by the development of genetic controllers that are able to titrate a given gene precisely. Classic molecular biology techniques such as transfection or lentiviral transduction for simple gene over expression will always be a mainstay. However, our field has recently adopted synthetic devices that incorporate linear and non-linear feedback in order to obtain more complex outputs from delivered DNA.

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