Compositions and methods for engineering cells
AssigneeLife Technologies Corporation
InventorsUma LAKSHMIPATHY, Bhaskar Thyagarajan, Jonathan Chesnut, Vasiliki Anest, Robert Bennett, Pauline Lieu, George Hanson, David Thompson, Lucas Chase, Gary Shipley, Elizabeth Wilson
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Abstract
The disclosure relates generally to genetic manipulation of stem and primary cells and to reprogramming of somatic cells, more specifically, human cells. In particular, compositions and methods are disclosed for the generation and maintenance of such engineered cells.
Claims23
An isolated nucleic acid molecule comprising
- (a) an OriP site,
- (b) a DNA segment encoding the EBNA1 gene wherein EBNA1 expression is inducible;
- (c) one or more att recombination sites containing one or more expression cassettes, each containing a promoter operably linked to a DNA sequence for which expression is desired, wherein the expression cassette encodes for a reprogramming gene selected from a group consisting of Oct4, Sox2, c-Myc and K1f4;
- Oct3/4, Nanog, Lin28, SSEA1, and TRA1-80;
- and
- (d) a DNA segment encoding at least one selectable marker.
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A kit comprising the pEBNA-DEST plasmid vector of SEQ. ID No.:
- 4, the pEP-EG plasmid vector of SEQ. ID No.:
- 5, or the pEP-hOG plasmid vector of SEQ. ID No.:
- 6.
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A method for reprogramming cells comprising:
- (i) introducing or expressing one or more small ds RNA molecules of SEQ ID NO:13-SEQ ID NO:48 into a cell;
- (ii) identifying whether said cell has been reprogrammed,
- wherein the small ds RNA molecules interacts with the promoter region of a reprogramming gene.
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Dependencies are inferred from the claim text.