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Published on: February 2, 2018
[Retrovirus vector expresses luciferase and TPO gene under the control of doxycycline in NIH3T3 cells]
1Institute of Hematology, People's Hospital, Beijing Medical University, Beijing 100044, China.
Objective:
To use the retrovirus vector Tet-On regulation system to control the expression of TPO and luciferase gene by doxycycline (Dox).
Methods:
Packing the recombinant Tet-On retrovirus, and infecting it to NIH3T3 cells to establish recombined Tet-On retrovirus cell lines, RevTet-On3T3. To construct the pRevTRETPO, the new cell line was transferred to PT67 cell to package the recombinant retrovirus which contained both the TRE and TPO. This virus was RevTet-On3T3 used to establish the two inserting recombinant retrovirus cell strain RevTet-On3T3TPO. Doxycycline was used to control the expression of TPO in RevTet-On3T3.
Results:
When Dox (5 mg/L) was added to the RevTet-On3T3 cells, a higher activity of luciferase was seen (mean, 5.9 x 10(4) RLU.S) compared with the mean activity (2.6 x 10(3) RLU.S) in the absence of Dox. When Dox was added to the RevTet-On3T3TPO cells, cell populations had high TPO expression (mean, 12.8 ng/ml) in 5 mg/L of Dox (On state), and a low TPO expression (mean, 0.56 ng/ml) in the absence of Dox (Off state).
Conclusion:
The RevTet-On gene expression system could modulate the expression of multiple genes by tetracycline and its derivatives, providing a useful tool for gene therapy.
