电脉冲介导荧光素酶基因转移高效率的实验研究

Electric pulse mediated high efficient luciferase gene transfer

目的 研究电脉冲介导的基因转移效率及其最佳基因转移电脉冲参数。方法 用微量注射器将pcDNA3-Luc质粒100μg注射入BALB/小鼠的股四头肌,5 min内在注射部位给予不同参数的电脉冲刺激,然后进行荧光素酶活性测定和组织病理切片。结果 电脉冲可明显增加荧光素酶基因的表达,电脉冲组的荧光素酶活性(7.26±3.72)×107 RLU/mg蛋白是单纯注射组(1.38±1.27)×104 RLU/mg蛋白的5 262倍(P=0.017)。组织病理检查发现电脉冲刺激无严重的肌肉组织损伤。电压100 V/cm,波宽60 ms,脉冲次数10次和频率0.75 Hz是最佳的电脉冲参数。结论 在最佳的电脉冲参数条件下,电脉冲介导的基因转移可获得较高的基因表达。

Objective To study gene transfer mediated by the electric pulse and optimize the parameters of electric pulse in vivo. Methods 100 μg plasmids of pcDNA3-Luc were injected into the quadriceps of BALB/c mice. Five minutes after DNA injection electric pulses with different parameters were given to the injection site. Two days after-ward, activity of luciferase was measured and the degree of muscle damage was analyzed by histological staining. Results The activity of luciferase in the electric pulse group [ (7. 26±3. 72) ×107 RLU/mg protein] was 5 262 folds higher than that in the simple injection group [ (1.38 ±1.27) ×104 RLU/mg protein] (P =0. 017). Histological analyses showed that in vivo electric pulse did not cause long-term severe muscle damage. When the electric pulse parameters were 100 V/cm,60 ms,10 pulses and 0.75 Hz, maximal gene expression was achieved. Conclusion Electri pulse,with optimal parameters, can increase gene expression obviously.