HSV1-tk基因重组逆转录病毒载体的构建与表达

The Construction of pN2AHSV1tk and Expression

通过ＤＮＡ重组技术，将ＨＳＶ１ｔｋ基因片段重组到含有ＨＳＶ１ｔｋ启动子的ｐＮ２Ａ型逆转录病毒载体中。选取正向克隆的逆转录病毒重组ＤＮＡ，以磷酸钙法转染ψ２，ＰＡ３１７包装细胞后，经Ｇ４１８筛选，抗性克隆细胞上清能成功地感染ＮＩＨ３Ｔ３，细胞系测定平均滴度为６．２×１０５ＣＦＵ／ｍＬ，最高可达１．５×１０６ＣＦＵ／ｍＬ。

The fulllength HSV1tk gene was inserted into retrovirus vector pN2A by DNA recombinant techniques. The plasmid pN2AHSV1tk was transfected into 2, PA317 packaging cell line by DNAcalcium phosphate coprecipitation. The G418 resistant clones were selected and their medium could infect NIH3T3 successfully and their average titer was 6.2105 CFU/mL. The highest one reached 1.5106 CFU/mL. The producer cell line will be used in gene therapy.