NEUROG2 is the gene encoding the neuronal transcription factor NGN2, which has been shown to accelerate neuronal differentiation when overexpressed in iPSCs. Via EBiSC, two iPS cell lines are available in which a Doxycycline (DOX) inducible construct to overexpress NEUROG2 either with (BIONi010-C-15) or without (BIONi010-C-13) a T2A-eGFP sequence into the safe-site locus AAVS1, have been inserted. Both lines remain pluripotent when kept in iPSC culture media, and efficiently differentiate into neurons after just three days of DOX treatment, without the need of extra neuronal differentiation factors.
Monoclonal subclones for BIONi010-C-13 and BIONi010-C-15 show a typical iPS cell morphology and in the absence of DOX continue to express the stem cell markers NANOG, TRA-1-60, OCT4, SSEA4 and SOX2. Additionally, they retain capacity for trilineage differentiation and are karyotypically normal. After a short period of DOX exposure, induction of the NGN2 insertion converts both iPSC lines to cells with a typical neuronal morphology. No pluripotent iPSCs remained in the differentiated cultures, confirming a pure, monoclonal population. After 14 days of differentiation, iNGN2 neurons expressed the basic neuronal marker Tuj1 and BIONi010-C-15 continues to express GFP.
DOX-iNGN2 neurons are a powerful tool to simplify reseearch into human neurons and EBiSC looks forward to sharing more research outputs using these iPSCs and derived neurons.
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Written by Rachel Steeg, May 2021
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