摘要
目的HVEM基因突变体腺病毒载体对大鼠混合淋巴细胞反应(MLR)的影响。方法RT-PCR获得人HVEM的cDNA,使其与LIGHT分子结合的表位点突变,保留与BTLA结合的关健位点。构建经修饰的HVEM基因的复制缺陷型腺病毒载体Adv-mHVEM-IRES2-EGFP并对其进行鉴定和转染效率测定。将该载体用于混合淋巴细胞反应中,观察对淋巴细胞增殖的影响。结果成功构建HVEM突变体和EGFP复制缺陷型腺病毒载体,经过测序与预期一致,体外培养细胞转染效率达到95%以上。该载体能够对大鼠MLR有明细的抑制作用。结论构建的腺病毒载体能够共表达目的基因的产物HVEM突变体和EGFP蛋白,对体外细胞具有高效转染能力。该载体能够对大鼠混合淋巴细胞反应有明显的抑制作用。
Objective To observe the efficacy of HVEM gene Adv vector in mixed lymphocyte reaction（MLR）. Methods HVEM cDNA bind domain of LIGHT was mutated by RT-PCR and PCR. Adenovirus expression vector Adv-mHVEM-IRES2-EGFP was constructed with mHVEM and EGFP by means o,f AD-Easy system. The vector was identified and evaluated the efficiency of transfection. The adenovirus vector was used in mixed lymphocyte reaction（MLR）. Results HVEM and EGFP were expressed in the culture cells transfected by the vector with a transfection efficiency over 95 %. MRL could be inhibited by the adenovirus vector. Conclusion The reeombinate vector Adv-mHVEM-IRES2-EGFP could express the protein of mHVEM and EGFP in cell line with high transfection efficiency. MRL could be inhibited by the adenovirus vector.
出处
《重庆医学》
CAS
CSCD
2006年第15期1366-1368,共3页
Chongqing medicine
基金
国家自然科学基金资助项目(30571863)
