**4.1 Materials**


8. TrypLE Express (Invitrogen).

100 Non-Viral Gene Therapy

section, we describe one example of retroviral transduction using our mechanical agitation

Fig. 4. A Typical Example of Application of Machanical Agitation Method to Transduction of EGFP-Carrying Retrovirus (Park et al., 2009). (A) The representative FACS plots of rat mesenchymal stem cells after transduction with Retro-EGFP using the static method (left panel), mechanical agitation of viruses with adhered cells (middle panel) and simultaneous mechanical agitation of retroviruses with suspended cells (right panel). (B) Numerical representation of the transduction efficiencies of EGFP retrovirus under the static protocol versus the new agitation protocol. The transduction efficiency is defined as the percentage of cells expressing EGFP as measured using FACSCalibur. The mean percentage of GFPpositive cells is presented as the average of three independent transduction experiments

2. pCAG-EGFP expression vector (Addgene, USA) and pMSCVneo retroviral vector

4. Rat mesenchymal stem cells (isolated by flushing the femurs of two-month-old female

5. DMEM/ High glucose (Hyclone, USA) with 10% FBS (Hyclone), 2 mM L-glutamin (Invitrogen, USA), 100 μg/ml streptomycin, and 100 U/ml penicillin (Sigma, USA).

protocol.

(+/- SEM, n=3).

**4.1 Materials** 

(Clontech, USA).

1. Packaging cell line PT67 (Clontech).

3. Lipofectamine 2,000 (Invitrogen).

SD rats (Damool Bioscience Co., Korea).

6. Selection drug, G418 (Sigma) and Polybrene (Calbiochem, USA). 7. Rat mesenchymal stem cell characterization kit (Millipore,USA).

