摘要
从小鼠脾细胞提取总ＲＮＡ，通过ＲＴＰＣＲ得到含信号肽的小鼠γ干扰素（ｍＩＦＮγ）全长ｃＤＮＡ，经测序鉴定后将其克隆到含标记基因ＮｅｏＲ的逆转录病毒载体ｐＬＸＳＮ中，采用脂质体法将其导入包装细胞ＰＡ３１７，经过包装，用含ｍＩＦＮγ基因的缺陷型病毒上清感染小鼠肝癌细胞，经Ｇ４１８筛选建立起ｍＩＦＮγ基因修饰株。ＰＣＲ、ＲＴＰＣＲ、Ｓｏｕｔｈｅｒｎ及Ｎｏｒｔｈｅｒｎ杂交结果显示有ｍＩＦＮγ及标记基因的转入和表达。生物学活性测定也表明该基因修饰株细胞培养上清中有一定活性的ＩＦＮγ分泌。
The total cellular RNA was isolated from mouse spleen cells and the whole length mouse interferongamma(mIFN)cDNA containing signal peptide sequence was obtained using RTPCRAfter confirmation by sequencing,it was introduced into the mouse hepatoma cell line H22 by retroviral vectorThe correct integration and expression of mIFN gene were verified by molecular biology methods iePCR,Southern blot,RTPCR and Northern blotBy bioassay,the activity of IFN was demonstrated in the culture supernatant of this genemodified clone
出处
《上海免疫学杂志》
CSCD
北大核心
1999年第2期79-81,共3页
Shanghai Journal of Immunology
基金
国家"863"高科技
国家自然科学基金
