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Generation of human embryonic stem-like cells using intronic RNA

  • US 9,567,591 B2
  • Filed: 05/07/2008
  • Issued: 02/14/2017
  • Est. Priority Date: 05/15/2003
  • Status: Active Grant
First Claim
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1. A method for inducing genomic DNA demethylation and hence activating Oct3/4 and Sox2 expression in mammalian cells, comprising the steps of:

  • (a) constructing a recombinant SpRNAi-RGFP nucleic acid composition that contains at least an intronic RNA gene silencing effector comprising SEQ. ID. NO. 39, SEQ. ID. NO. 40, SEQ. ID. NO. 41, SEQ. ID. NO. 42, SEQ. ID. NO. 43, SEQ. ID. NO. 44, SEQ. ID. NO. 45 and SEQ. ID. NO. 46, wherein said intronic RNA gene silencing effector is further processed into mir-302a, mir-302b, mir-302c and mir-302d in mammalian cells;

    (b) introducing said recombinant SpRNAi-RGFP nucleic acid composition into a plurality of mammalian cells, wherein said plurality of mammalian cells generate a plurality of intronic RNA gene silencing effectors; and

    (c) enabling the plurality of intronic RNA gene silencing effectors to reach a level that is sufficient to induce genomic DNA demethylation and hence activate Oct3/4 and Sox2 expression, which consequently results in reprogramming the cells into a pluripotent state.

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