Single lentiviral vector system for induced pluripotent (iPS) stem cells derivation

摘要

The present invention is based on the discovery that a single lentiviral vector expressing multiple individual transcription factor proteins from a single multi-cistronic mRNA can reprogram a fibroblast cell to a stem cell-like cell. These reprogrammed induced pluripotent stem (iPS) cells are pluripotent. Additions of the Cre-LoxP sequences into the single lentiviral vector facilitate excision of the vector after reprogramming in achieved. Addition of a maker gene into the single lentiviral vector facilitates detection of the presence of the vector in an iPS. The invention provides compositions and methods of producing iPS cells using a single multi-cistronic lentiviral vector.

主权项

1. A lentiviral vector particle wherein the vector particle comprises a nucleic acid sequence comprising a sequence encoding:
a. a first gene; b. a second gene; c. a third gene; d. fourth gene, wherein the four genes are Oct4, Sox2, Klf4 and cMyc, or Oct4, Sox2, Lin28, and Nanog; e. a first ‘self-cleaving’ 2A peptide; f. second ‘self-cleaving’ 2A peptide; and g. an internal ribosome entry site (IRES); wherein the first 2A ‘self-cleaving’ peptide is positioned between at least the first and second genes and the second ‘self-cleaving’ 2A peptide is positioned between the third and fourth genes, wherein the first 2A ‘self-cleaving’ peptide and second ‘self-cleaving’ 2A peptide are different, wherein the nucleic acid sequence is operably linked to a promoter, wherein the IRES is between the second and third genes, and wherein the sequences encoding the first, second, third and fourth genes, first and second ‘self-cleaving’ 2A peptides, and the IRES are transcribed from the promoter as a multi-cistronic RNA.