Data for: Plasmid BMP-2–Embedded Gelatin Sponge As A Gene-Activated Matrix for Preosteoblast Differentiation

Supplementary Figure 1
To allow endocytosis of DNA complexes into the cells, 5 × 104 cells and TransIT-2020/pEGFP-C1 complexes were mixed for 5, 15, 30, or 60 min (n = 3). Flow cytometry analysis was performed to evaluate GFP expression at these time points using FACS Calibur flow cytometer (Becton Dickinson, Heidelberg, Germany). Cells were gated at 1 × 104 cells/sample in FL1 channel setting, and BMP-2 transfection samples under the same condition at each time point were used as controls(Figure S1A). Transfection efficiencies of these four mixing time points were compared using Becton Dickinson Cell Quest software(Figure S1B). We found that duration of 30 min was the optimal mixing time, as the average GFP expression (9.92%) at 30 min was the highest compared to that at the other three time points.

Live and dead assay :
The scaffolds were stained using a Live/Dead Cytotoxicity/Viability kit (Invitrogen, Carlsbad, CA)from day 7 to day 28 .The live cells ,dead cells and merge images were all included.

Figure 5 ALP supplementary data : all ALP activity corresponding to Figure 5
Figure 6 ARS supplementary data : all ARS activity corresponding to Figure 6
Figure 7 BMP-2 supplementary data: all BMP-2 expression corresponding to Figure 7

补充图1
为使DNA复合物顺利内吞进入细胞，将5×10⁴个细胞与TransIT-2020/pEGFP-C1复合物分别孵育5、15、30或60分钟（n=3）。采用FACSCalibur流式细胞仪（Becton Dickinson，德国海德堡）对各时间点的GFP表达水平进行流式细胞术分析。每个样本以1×10⁴个细胞在FL1通道设门，并以相同条件下各时间点的BMP-2转染样本作为对照（图S1A）。使用Becton Dickinson Cell Quest软件比较四个孵育时间点的转染效率（图S1B）。结果显示，30分钟为最优孵育时长，该时间点的平均GFP表达率（9.92%）显著高于其余三个时间点。

死活细胞染色实验：
分别于第7天至第28天，使用Invitrogen（美国加利福尼亚州卡尔斯巴德）生产的Live/Dead细胞毒性/活性检测试剂盒对支架进行染色，实验结果包含活细胞图像、死细胞图像及合并叠加图像。

图5 ALP补充数据：对应图5的全部碱性磷酸酶（ALP）活性数据
图6 ARS补充数据：对应图6的全部茜素红S（ARS）活性数据
图7 BMP-2补充数据：对应图7的全部骨形态发生蛋白2（BMP-2）表达数据