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20 February 2017 DDX4-EGFP transgenic rat model for the study of germline development and spermatogenesis
Kathrin Gassei, Yi Sheng, Adetunji Fayomi, Payal Mital, Meena Sukhwani, Chih-Cheng Lin, Karen A. Peters, Andrew Althouse, Hanna Valli, Kyle E. Orwig
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Abstract

Spermatogonial stem cells (SSC) are essential for spermatogenesis and male fertility. In addition, these adult tissue stem cells can be used as vehicles for germline modification in animal models and may have application for treating male infertility. To facilitate the investigation of SSCs and germ lineage development in rats, we generated a DEAD-box helicase 4 (DDX4) (VASA) promoter-enhanced green fluorescent protein (EGFP) reporter transgenic rat. Quantitative realtime polymerase chain reaction and immunofluorescence confirmed that EGFP was expressed in the germ cells of the ovaries and testes and was absent in somatic cells and tissues. Germ cell transplantation demonstrated that the EGFP-positive germ cell population from DDX4-EGFP rat testes contained SSCs capable of establishing spermatogenesis in experimentally infertile mouse recipient testes. EGFP-positive germ cells could be easily isolated by fluorescence-activated cells sorting, while simultaneously removing testicular somatic cells from DDX4-EGFP rat pup testes. The EGFP-positive fraction provided an optimal cell suspension to establish rat SSC cultures that maintained long-term expression of zinc finger and BTB domain containing 16 (ZBTB16) and spaltlike transcription factor 4 (SALL4), two markers ofmouse SSCs that are conserved in rats. The novel DDX4-EGFP germ cell reporter rat described here combined with previously described GCS-EGFP rats, rat SSC culture and gene editing tools will improve the utility of the rat model for studying stem cells and germ lineage development.

Summary Sentence

A DDX4 (VASA)-EGFP transgenic rat was produced to facilitate investigations of stem cells and germ lineage development.

© The Authors 2017. Published by Oxford University Press on behalf of Society for the Study of Reproduction. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.
Kathrin Gassei, Yi Sheng, Adetunji Fayomi, Payal Mital, Meena Sukhwani, Chih-Cheng Lin, Karen A. Peters, Andrew Althouse, Hanna Valli, and Kyle E. Orwig "DDX4-EGFP transgenic rat model for the study of germline development and spermatogenesis," Biology of Reproduction 96(3), 707-719, (20 February 2017). https://doi.org/10.1095/biolreprod.116.142828
Received: 21 June 2016; Accepted: 30 January 2017; Published: 20 February 2017
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