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目的 研究电脉冲介导的基因转移效率及其最佳的基因转移的电脉冲参数。方法 用微量注射器将pcD2 /LacZ质粒 10 μg ,在昆明小鼠的股四头肌注射 ,1～ 2min内在注射部位给予不同参数的电脉冲刺激 (不同的电脉冲参数每组 10只小鼠 ) ,然后进行 β 半乳糖苷酶活性的测定或酶组织化学染色。同时设空白对照组及单纯注射组。结果 电脉冲可明显增加LacZ基因的表达 ,电脉冲组的β 半乳糖苷酶活性 (131.6U/mg± 86 .5U/mg蛋白 )是单纯注射组 (4 .9U/mg± 1.0U/mg蛋白 )的 30倍(P <0 .0 5 )。组织化学染色结果表明电脉冲组肌肉组织中 β 半乳糖苷酶蓝色颗粒的数目和染色的程度均明显高于单纯肌肉注射组。当电脉冲参数电压 2 0 0V/cm ,波宽 40ms,脉冲次数 6次和频率 1Hz时 ,可获得最高的基因表达效率。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 the electrical pulse-mediated gene transfer efficiency and its optimal electrical pulse parameters for gene transfer. Methods 10 μg of pcD2 / LacZ plasmid was injected into femoral quadriceps of Kunming mice by microinjector. Electro-pulsed stimulation with different parameters was given at injection site within 1 ~ 2 minutes (10 different mice with different electrical pulse parameters) The determination of beta-galactosidase activity or enzymatic histochemical staining is then performed. At the same time set blank control group and simple injection group. Results The electric pulse significantly increased the expression of LacZ gene. The β-galactosidase activity (131.6U / mg ± 86.5U / mg protein) in the electric pulse group was significantly higher than that in the pure injection group (4.9U / mg ± 1.0U / mg protein ) (P <0.05). Histochemical staining results showed that the number and staining degree of β-galactosidase blue granules in muscle tissue of electric pulse group were significantly higher than that of simple intramuscular injection group. The highest gene expression efficiency was obtained when the parameters of electrical pulse voltage were 200 V / cm, wave width 40 ms, pulse number 6 times and frequency 1 Hz. Conclusion Electrical pulse-mediated gene transfer can achieve higher gene expression under the optimal electrical pulse parameters.