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. 2020 Jul 9;10:11393. doi: 10.1038/s41598-020-68257-y

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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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dCas9-KRAB represses PTEN and enhances neurite outgrowth in rat PC-12 cell line differentiated to neural phenotype. Rattus norvegicus PC-12 cells were transduced with dCas9-KRAB and PTEN gRNA D, four shRNAs targeting PTEN, or EGFP control, either before (A) or after (B–E) being differentiated to a neural phenotype with nerve growth factor (NGF). (A,B) Pten mRNA expression in qRT-PCR when virus was delivered to undifferentiated PC-12 cells (A) or to PC-12 cells that had already been stimulated with NGF for 48 h (B), shown as fold change relative to EGFP control, *p < 0.05 (Kruskall-Wallis test with Dunn’s multiple comparisons test to compare each condition to EGFP control), n = 3. (C) Representative images of differentiated PC-12 cells expressing EGFP, with phase contrast illustrating neurite length. Scale bar = 500 µm. (D) The percentage of transduced cells extending 1 or more neurites equal to or greater than the diameter of the cell body. (E) Total neurite length per differentiating, transduced cell. **p < 0.01 (One-way ANOVA with Dunnett’s multiple comparisons test to compare each condition to EGFP control), n = 3, error bars show SEM.