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Updated: Jun 17, 2026
Preparation of rAAV9 to Overexpress or Knockdown Genes in Mouse Hearts
Published on: December 17, 2016
[Recombinant adeno-associated virus serotype 9 transfection of rats H9C2 cells in vitro]
Xia Gao1, Yi-tong Ma, Yi-ning Yang
1Department of Cardiology, The First Affiliated Hospital of Xinjiang Medical University, Urumqi 830054, China. gaoxia999@gmail.com
Aim:
To evaluate the transfection efficiency using recombinant adeno-associated virus Serotype 9 mediated enhanced green fluorescent protein (rAAV9- EGFP) to rats H9C2 cells and the impact on growth of H9C2 cells.
Methods:
rAAV9-EGFP was transfected into H9C2 cells at different multiplicities of infection (MOI=1 x 10(5), 1 x 10(6), 1 x 10(7)). EGFP expression in the cells was observed under inverted fluorescence microscope, and the EGFP-positive cell percentage determined by flow cytometry. Alamar Blue assay was used to assess the proliferation of the transfected cells.
Results:
The cells with rAAV9-EGFP transfection at MOI of 1 x 10(6) and 1 x 10(7) began to exhibit EGFP expression 1 days after transfection and the cells transfection at MOI of 1 x 10(5) began to exhibit EGFP expression 2 days after transfection. the fluorescence intensity increased with the MOI used for transfection. EGFP expression reached the maximum on day 4, at the point of which the transduction efficiency of rAAV9-EGFP in H9C2 cells was (14.1+/-0.2)%, (35.1+/-4.8)% and (56.8+/-0.1)%. Corresponding to MOIs of 1 x 10(5), 1 x 10(6) and 1 x 10(7), respectively. Alamar Blue assay did not reveal significant difference in the absorbance between the transfected cells and the control cells after transfection.
Conclusion:
rAAV9-EGFP gene can be stably and efficiently expressed in H9C2 cells without causing cell growth inhibition.This study played foundation for further research.
