Construction of the Recombinant Lentiviral Vector Containing Mouse Tox3 Gene and Its Expression in Primary Granulosa Cells

Objective: To construct the recombinant lentiviral vector containing mouse Tox3 gene, and to check its expression in primary granulosa cells. Methods: The full length of Tox3 fragment was amplified by PCR, and subcloned into the lentiviral vector pCDH-MCS-T2A-copGFP-MSCV. The resultant lentivirus were confirmed by PCR, restriction enzyme digestion and DNA sequencing. The recombinant lentivirus(LV-Tox3-Flag) were produced from HEK293FT by a transient co-transfection with psPAX2 and pMD2.G. Primary granulosa cells were infected by LV-Tox3-Flag lentivirus,and the expression of TOX3 was confirmed by RTPCR and Western blot. Results: The recombinant lentiviral vector carried the Tox3 gene was successfully constructed. RT-PCR and Western blot analysis revealed that the Tox3 gene can be correctly transcript and translated in the granulosa cells infected by LV-Tox3. Conclusions: The recombinant LV-Tox3-Flag was successfully constructed. It will be used to transfect target cells in our future study.