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Updated: Jun 10, 2026
A Seamless Cloning Approach for Porcine Reproductive and Respiratory Syndrome Virus Expression Vector Construction
Published on: May 17, 2024
[Cloning of hsa-miR-148a and construction of its retroviral expression vector]
Xue-hu Xu1, Tian-yi Quan, Wei-xia Zeng
1Department of General Surgery, Third Affiliated Hospital of Guangzhou Medical College, Guangzhou 510150, China. maxtiger@126.com
Objective:
To clone hsa-miR-148a and construct its retroviral expression vector.
Methods:
The pre-miR-148a amplified by PCR was inserted to pMSCV to construct the recombinant retroviral expression plasmid pMSCV-miR-148a, which was confirmed by restriction endonuclease analysis and DNA sequencing. The retroviral expression vector pMSCV-miR-148a and PIK packaging plasmid were cotransfected into 293FT packaging cells by calcium phosphate-mediated transfection to produce the retrovirus, and the retrovirus titer was measured by infection of NIH3T3 cells.
Results:
Restriction enzyme digestion and DNA sequencing demonstrated that the retroviral vector pMSCV-miR-148a was constructed successfully, and the virus titer was 5x10(8) CFU/ml after infection of NIH3T3 cells.
Conclusion:
The successful construction of the retroviral expression vector MSCV-miR-148a allows the production of high-titer retrovirus to facilitate further study of the molecular functions of miR-148a.
