A vector-based system for the differentiation of mouse embryonic stem cells toward germ-line cells

Ebrahimzadeh-Vesal, Reza and hokrgozar, Mohammad Ali and Nayernia, Karim and Teimoori-Toolabi, Ladan and Miryounesi, Mohammad and Nourashrafeddin, Seyedmehdi and Ranji, Najmeh and Modarressi, Mohammad Hosein (2014) A vector-based system for the differentiation of mouse embryonic stem cells toward germ-line cells. Iranian Journal of Basic Medical Sciences, 17 (8). pp. 566-570.

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Official URL: http://ijbms.mums.ac.ir/article_3190.html

Abstract

Objective(s):To culture thein vitro mouse embryonic stem cells (mESCs) and to direct their differentiation to germ-line cells; in present study we used a vector backbone containing the fusion construct Stra8-EGFP to select differentiated ES cells that entered meiosis. Retinoic acid was used to differentiate embryonic stem cells to germ cells. Materials and Methods: A fragment of Stra8 gene promoter (-1400 to +7) was inserted in ScaI/HindIII multiple cloning site of pEGFP-1 vector. Theelectroporationwas done on embryonic stem cells and positive colonies were selected as puromycin-resistant after three weeks of treatment with puromycin. All-trans retinoic acid (RA) was used for differentiation of mESCs at final concentration of 10-5M. The expression of protamine 1 (Prm1) gene was checked as post meiotic marker in differentiated mESCs after 5, 10, 15, 21 and 30 days after RA induction. Results: The PCR amplification by specific primers for Stra8-EGFP fusion gene was detected in DNA sample from mESCs after electroporation and puromycin treatment. GFP-positive mESC colonies were observed after 72 hr RA induction. The protamine 1 gene was expressed after 21 days of RA induction. Conclusion: In this study, we demonstrated the in vitro generation of mouse embryonic stem cells to germ cells by using a backbone vector containing the fusion gene Stra8-EGFP. The Stra8 gene is a retinoic acid-responsive protein and is able to regulate meiotic initiation.

Item Type: Article
Subjects: QV pharmacology
Divisions: Journals > Iranian J Basic Medical Sciences
Depositing User: ijbms ijbms
Date Deposited: 07 Oct 2017 15:45
Last Modified: 07 Oct 2017 15:45
URI: http://eprints.mums.ac.ir/id/eprint/7794

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