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Cytotechnology

, Volume 70, Issue 1, pp 45–53 | Cite as

Markers associated with neuron-specific Ube3a imprinting during neuronal differentiation of mouse embryonic stem cells

  • Masamitsu Eitoku
  • Hidemasa Kato
  • Narufumi Suganuma
  • Hidenori Kiyosawa
Method in Cell Science

Abstract

Understanding gene expression in the brain requires allele-specific transcriptome analysis because of the presence of neuron-specific imprinted genes, which are expressed in a neuron-specific and parent-of-origin-specific manner. Ube3a is a neuron-specific imprinted gene with an expression pattern that changes from biallelic to maternal only (Ube3a imprinting) during differentiation. Because Ube3a imprinting occurs only in neurons, it has the potential to be a marker to assess the quality of neurons produced by in vitro neuronal differentiation of embryonic stem cells (ESCs). For the analysis of Ube3a imprinting, genetic polymorphisms between the two alleles are necessary to identify the parental origin of each. However, ESCs derived from commonly used inbred mouse strains have no genetic polymorphisms. To overcome this problem, we examined 10 markers of neurogenesis to determine whether they were associated with Ube3a imprinting. We measured the relative expression levels of these 10 gene markers and assessed the Ube3a imprinting status of 54 neuron samples differentiated under various in vitro conditions. Then we divided the samples into two groups depending on their Ube3a imprinting status and selected markers statistically associated with Ube3a imprinting. The identified markers included the antisense noncoding transcript of Ube3a and a mature neuron marker Mtap2, consistent with the markers we used empirically in our previous study to assess the quality of differentiated neurons. These findings provide new quality control criteria for differentiated neurons, and could also be applied to human ESCs and induced pluripotent stem cells.

Keywords

Ube3a imprinting Embryonic stem cell In vitro neuronal differentiation Mus musculus molossinus Mtap2 Noncoding RNA SNP ratio quantification 

Notes

Acknowledgements

This work was financially supported by a Japan Society for the Promotion of Science (JSPS) Grant-in-Aid for Scientific Research (KAKENHI 24115703 and 26290063) and a JSPS Grant-in-Aid for Young Scientists (KAKENHI 16K19273). The authors thank Dr. K. Araki for embryonic stem cell lines and Ms. H. L. Le, M. Okabayashi, and Mr. H. Makino for technical assistance.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

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Copyright information

© Springer Science+Business Media B.V. 2017

Authors and Affiliations

  1. 1.Department of Environmental Medicine, Kochi Medical SchoolKochi UniversityOko-Cho Kohasu NankokuJapan
  2. 2.Division of Functional Histology, Department of Functional Biomedicine, Graduate School of MedicineEhime UniversityToonJapan
  3. 3.Department of Life ScienceChiba Institute of TechnologyNarashino, ChibaJapan

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