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. 2020 Jun 5;10:9262. doi: 10.1038/s41598-020-66266-5

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© The Author(s) 2020

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Differentiation of CMTX6 patient-derived and control iPSCs into motor neurons. (A) Timeline for spinal cord motor neuron differentiation from iPSC following a dual SMAD inhibition protocol. Diagram shows media used and factors added throughout the process. Cells were grown in suspension (3D) between days 18 and 26. Abbreviations: DM (Dorsomorphin); SB (SB431542); RA (Retinoic acid); SAG (Smoothened agonist). GDNF (Glial cell line-derived neurotrophic factor); CNTF (Human Ciliary neurotrophic factor); BDNF (Brain-derived neurotrophic factor); phosphorylated 7-ethyl-10-hydroxycamptothecin (SN38-P). (B) Differentiated motor neurons show a robust expression of ISL1, HB9 and NEFL markers at DIV 32. Motor neurons organise into clusters of cell bodies at longer maturation times (DIV 37, DIV 42) with connecting axons (TUBB3) occupying larger areas of the culture.