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Construction of Fzd6Q152E mice through CRISPR/Cas9 technology and their reproduction and identification

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Abstract

Background

The CRISPR/Cas9 system is widely used for genome editing in human, rat and mouse cells. In this study, we established Fzd6 mutant mice using CRISPR/Cas9 technology, and obtained Fzd6 homozygous mutant (Fzd6Q152E) mice through breeding. Fzd6 plays a role in depression, but there are few related reports. We used this model to investigate the mechanism of Fzd6 involved in depression, and build a solid foundation for subsequent in-depth studies.

Methods and Results

The target of Fzd6 mutation was obtained by CRISPR/Cas9 technology and hippocampal tissue was collected for Nissl staining and histological analysis. Blood was collected for enzyme linked immunosorbent assay (ELISA); The gene expression of Fzd6 and the related genes expression in wnt pathway was quantified by quantitative real-time PCR (qRT-PCR), and then expression of Fzd6 and proteins in the Wnt pathway were identified by western blotting. ELISA results showed that the expression levels of brain derived neurotrophic factor (BDNF), 5-hydroxytryptamine (5-HT), and Noradrenaline (NE) in serum were significantly decreased in Fzd6Q152E mice, whereas the mRNA expression of Lrp5, Lrp6, and Dkk2 is increased. The western blotting revealed that the expression of Fzd6 and Lrp6 is decreased, although the expression of Dkk2 and Gsk-3β increased.

Conclusion

Our study successfully established homozygous Fzd6 mutant mice model. The relationship between Fzd6-Wnt and depression was preliminarily clarified, which provides an ideal animal model for subsequent research on diseases induced by the Fzd6 mutation.

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Acknowledgements

This work was funded and supported by the scientific research project of Shanxi Provincial Health Commission (2021031) and the Provincial Key Laboratory of Biological Psychiatry, the 2020 "Four Batches" Science and Technology Innovation Program (2020SYS10). We would like to thank Editage (www.editage.cn) for English language editing.

Funding

This work was funded and supported by the scientific research project of Shanxi Provincial Health Commission (2021031) and the Provincial Key Laboratory of Biological Psychiatry, the 2020 "Four Batches" Science and Technology Innovation Program (Grant No. 2020SYS10).

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Author notes

  1. Xiaoru Yan and Junting Yang have contributed equally to this work.

Authors and Affiliations

  1. School of Mental Health, Shanxi Medical University, South Shifang Street 55, Taiyuan, 030001, China

    Xiaoru Yan, Liang Wang, Yang Qi & Guohua Song

  2. Laboratory Animal Center, Shanxi Key Laboratory of Experimental Animal Science and Human Disease Animal Model, Shanxi Medical University, Road Xinjian 56, Taiyuan, 030001, Shanxi, China

    Xiaoru Yan, Junting Yang, Lanfei Xiao, Jiping Gao, Guoqiang Xu & Guohua Song

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  1. Xiaoru Yan
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  2. Junting Yang
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Contributions

XY and GS: contributed to the study conception, The first draft of the manuscript was written by XY and all authors commented on previous versions of the manuscript. XY, JY, LX, GX, LW and YQ: performed animal experiments, molecular experiments and data analysis.

Corresponding author

Correspondence to Guohua Song.

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All authors declare that they have no conflicts of interest.

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The protocol was approved by the Institutional Animal Care and Use Committee of Shanxi Medical University (SYDL2021012).

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Yan, X., Yang, J., Xiao, L. et al. Construction of Fzd6Q152E mice through CRISPR/Cas9 technology and their reproduction and identification. Mol Biol Rep 49, 9575–9584 (2022). https://doi.org/10.1007/s11033-022-07848-6

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  • DOI: https://doi.org/10.1007/s11033-022-07848-6

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