Adeno-X Tet-On系统调控LacZ基因在雪旺细胞中表达的研究  

Inducible expression of LacZ gene in Schwann cells by Adeno-X Tet-On system

作　　者：郝伟[1] 朱锦宇[1] 朱庆生[1] 李志全[1] 杨安钢[2] 

机构地区：[1]第四军医大学西京医院骨科,西安710033 [2]第四军医大学基础部免疫学教研室

出　　处：《山西医科大学学报》2004年第2期118-121,共4页Journal of Shanxi Medical University

基　　金：国家自然科学基金资助项目 ( 3 0 2 713 15 )

摘　　要：目的　应用Adeno XTet On系统感染雪旺细胞 ,通过四环素衍生物强力霉素 (doxcycline ,Dox)调控LacZ基因产物 (βgal)的表达。方法　①取生后 5～ 6dSD仔鼠坐骨神经进行雪旺细胞培养 ,并进行形态学及S 10 0蛋白相关抗原免疫组织化学染色鉴定。②将Adeno XTet OnVirusStock与Adeno XTRE βgalVirusStock分别感染HEK2 93细胞 ,收获病毒并作滴度测定。③将扩增后的病毒感染雪旺细胞并绘制Dox调控下βgal诱导表达曲线。结果　①由体外培养获得的雪旺细胞纯度可达 90 %以上并且状态良好 ,免疫组织化学染色阳性结果明显。②扩增后的病毒滴度较高 ,分别达 1.2 6× 10 9、1.99× 10 9pfu/ml。　结论　将病毒感染雪旺细胞后 ,经检测发现DOX对Objective To infect Schwann cells by Adeno-X Tet-On system and observe the inducible expression of LacZ gene under the regulation of doxcycline. Methods ①Five to six-day-old Sprague-Dawley rats were sacrificed by decapitation and the sciatic nerves were immediately removed aseptically to get Schwann cells. After the cells had been cultured and purified, those were identified by indirect immunocytochemistry with mouse anti-S100 protein antibody McAb.②Adeno-X Tet-On virus stock and Adeno-X TRE-βgal virus stock were amplified by infecting HEK293 cells respectively. When CPE was evident,the viruses were isolatedand viral titer were determined.③Schwann cells were infected with amplified viruses and the data of inducible expression of LacZ gene under the regulation of doxcycline were ploted. Results ①The purity of Schwann cells was more than90% and in a good state. ②The titer of amplified viruses was 1.26×10 9 and1.99×10 9 pfu/ml respectively. Conclusion After Schwann cells were infected with Adeno-X Tet-On virus and Adeno-X TRE-β gal virus, it was found that the concerntration change of Dox had a strong influence on the expression of LacZ gene.

关 键 词：Adeno-X Tet-On系统 LACZ基因 雪旺细胞 免疫组织化学 周围神经损伤 重组腺病毒载体 

分 类 号：R741[医药卫生—神经病学与精神病学]