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Visualization of Turbot (Scophthalmus maximus) Primordial Germ Cells in vivo Using Fluorescent Protein Mediated by the 3′ Untranslated Region of nanos3 or vasa Gene

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Abstract

Primordial germ cells (PGCs) as the precursors of germ cells are responsible for transmitting genetic information to the next generation. Visualization of teleost PGCs in vivo is essential to research the origination and development of germ cells and facilitate further manipulation on PGCs isolation, cryopreservation, and surrogate breeding. In this study, artificially synthesized mRNAs constructed by fusing fluorescent protein coding region to the 3′ untranslated region (3′UTR) of nanos3 or vasa (mCherry-Smnanos3 3′UTR or mCherry-Smvasa 3′UTR mRNA) were injected into turbot (Scophthalmus maximus) fertilized eggs for tracing PGCs. The results demonstrated that the fluorescent PGCs differentiated from somatic cells and aligned on both sides of the trunk at the early segmentation period, then migrated and located at the dorsal part of the gut where the gonad would form. In the same way, we also found that the zebrafish (Danio rerio) vasa 3′UTR could trace turbot PGCs, while the vasa 3′UTR s of marine medaka (Oryzias melastigma) and red seabream (Pagrus major) failed, although they could label the marine medaka PGCs. In addition, through comparative analysis, we discovered that some potential sequence elements in the3 ′UTRs of nanos3 and vasa, such as GCACs, 62-bp U-rich regions and nucleotide 187–218 regions might be involved in PGCs stabilization. The results of this study provided an efficient, rapid, and specific non-transgenic approach for visualizing PGCs of economical marine fish in vivo.

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Funding

This research was funded by National Natural Science Foundation of China (31572602), National Key Research and Development Program (2018YFD0901205, 2018YFD0901204), the Marine S&T Fund of Shandong Province for Pilot National Laboratory for Marine Science and Technology(Qingdao)(2018SDKJ0302-4, 2018SDKJ0502-2),China Agriculture Research System (CARS-47), STS project (KFZD-SW-106, 2017T3017), Shandong Province Key Research and Invention Program (2017CXGC010K), the National Infrastructure of Fishery Germplasm Resource (2019DKA30470).

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Authors and Affiliations

  1. CAS Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, 7 Nanhai Road, Qingdao, 266071, P. R. China

    Li Zhou, Xueying Wang, Qinghua Liu, Shihong Xu, Haixia Zhao, Mingming Han, Yunong Wang & Jun Li

  2. Laboratory for Marine Biology and Biotechnology, Qingdao National Laboratory for Marine Science and Technology, 7 Nanhai Road, Qingdao, 266071, P. R. China

    Li Zhou, Xueying Wang, Qinghua Liu, Shihong Xu, Haixia Zhao, Mingming Han, Yunong Wang & Jun Li

  3. Center for Ocean Mega-Science, Chinese Academy of Sciences, 7 Nanhai Road, Qingdao, 266071, P. R. China

    Li Zhou, Xueying Wang, Qinghua Liu, Shihong Xu, Haixia Zhao, Mingming Han, Yunong Wang & Jun Li

  4. University of Chinese Academy of Sciences, Beijing, China

    Li Zhou, Haixia Zhao & Yunong Wang

  5. Weihai Shenghang Aquatic Product Science and Technology Co. Ltd., Weihai, 264200, China

    Zongcheng Song

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  1. Li Zhou
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  8. Zongcheng Song
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  9. Jun Li
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Contributions

This study was conceived and designed by Li Zhou, Qinghua Liu, Xueying Wang, and Jun Li; Li Zhou and Xueying Wang contributed experimental work; Li Zhou and Qinghua Liu contributed the manuscript writing and revision, respectively; Shihong Xu, Haixia Zhao, Mingming Han, and Yunong Wang contributed experimental materials. All authors read and approve the final manuscript.

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Correspondence to Qinghua Liu or Jun Li.

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All experiments were performed in accordance with the relevant national and international guidelines and approved by the Institutional Animal Care and Use Committee, Institute of Oceanology, Chinese Academy of Sciences.

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Zhou, L., Wang, X., Liu, Q. et al. Visualization of Turbot (Scophthalmus maximus) Primordial Germ Cells in vivo Using Fluorescent Protein Mediated by the 3′ Untranslated Region of nanos3 or vasa Gene. Mar Biotechnol 21, 671–682 (2019). https://doi.org/10.1007/s10126-019-09911-z

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