期刊论文

Establishment of interferon membrane protein IFITM1 knockdown cells and investigation of its effect on the replication of PRV,PCV2 and TGEV

中文

中国兽医科学

1673-4696

2020

50

5

603-609

10.16656/j.issn.1673-4696.2020.0082

本校为第一机构

为构建稳定干扰(IFITM1)基因表达的猪空肠上皮细胞系J2-KD (knockdown,KD),并研究干扰IFITM1表达后对伪狂犬病病毒(PRV)、猪圆环病毒2型(PCV2)、猪传染性胃肠炎病毒(TGEV)感染的影响.应用基因克隆技术,构建pLKO.1-EGFP-Puro-IFITM1重组载体,与慢病毒包装质粒共转染293FT细胞,产生绿色荧光蛋白标记的表达IFITM shRNA的慢病毒,48 h后收集病毒上清.用收获的慢病毒感染猪小肠上皮细胞(J2细胞)后,经过嘌呤霉素筛选及细胞有限稀释法,通过RT-PCR鉴定后,获得稳定干扰IFITM1基因的细胞系,并由MTT法验证干扰IFITM1表达对J2细胞生长无影响,将成功构建的干扰IFITM1表达的J2稳定细胞系命名为J2-KD.分别用PCV2...

To establish a J2-KD(knockdown)cell line stably expressing interfered IFITM1 and study the effect of interference with IFITM1 gene on the infection of PCV2,PRV and TGEV.Gene cloning techniques were used to constructed pLKO.1-EGFP-Puro-IFITM1 recombinant vector,which was co-transfected into 293FT cells with lentiviral packaging plasmids psPAX2 and pMD2.G to produce green fluorescent protein labeled lentiviruses expression IFITM1shRNA,the viral supernatant was collected at 48 hours after post transfection.J2 cells were infected with the harvested lentiviruses,screened by puromycin and cloned via...