CAJ | 학술논문

目的探讨RNA干扰对单纯疱疹病毒2型(HSV2)ICP4基因表达和HSV2 DNA复制的抑制作用.方法化学合成靶向作用于HSV2 ICP4的四对siRNA和阴性对照siRNA,分别命名为siRNA-1,siRNA-2,siRNA-3,siRNA-4及siRNA-N,转染Vero细胞,过夜后用HSV2 HG52标准毒株感染上述Veto细胞,1、2、3、4d和5d收集Vero细胞和上清液,荧光定量RT-PCR检测ICP4 mRNA的表达水平,荧光定量PCR检测HSV2 DNA拷贝数,Western-Blot检测ICP4蛋白表达水平.结果与阴性对照相比,四对siRNA对HSV2 ICP4 mRNA和蛋白均有不同程度的抑制作用,以siRNA-2抑制作用最强,并且均可显著降低HSV2 DNA拷贝数.结论靶向作用于HSV2 ICP4的siRNA可显著抑制ICP4mRNA及蛋白表达和HSV2 DNA拷贝数,提示siRNA可通过抑制HSV2 ICP4基因表达而抑制HSV2的复制.
목적 탐토RNA간우대단순포진병독2형(HSV2)ICP4기인표체화HSV2 DNA복제적억제작용.방법 화학합성파향작용우HSV2 ICP4적사대siRNA화음성대조siRNA,분별명명위siRNA-1,siRNA-2,siRNA-3,siRNA-4급siRNA-N,전염Vero세포,과야후용HSV2 HG52표준독주감염상술Veto세포,1、2、3、4d화5d수집Vero세포화상청액,형광정량RT-PCR검측ICP4 mRNA적표체수평,형광정량PCR검측HSV2 DNA고패수,Western-Blot검측ICP4단백표체수평.결과 여음성대조상비,사대siRNA대HSV2 ICP4 mRNA화단백균유불동정도적억제작용,이siRNA-2억제작용최강,병차균가현저강저HSV2 DNA고패수.결론 파향작용우HSV2 ICP4적siRNA가현저억제ICP4mRNA급단백표체화HSV2 DNA고패수,제시siRNA가통과억제HSV2 ICP4기인표체이억제HSV2적복제.
Objective To explore the inhibition effect of RNA interference on the IcP4 expression and DNA replication of herpes simplex virus type 2(HSV2).Methods Four pairs of siRNA targeted to HSV2 ICP4 gene and negative control siRNA were synthetized by chemieal method.named as siRNA-1,siRNA-2,siRNA-3,siRNA-4 and siRNA-N respeeticely.HSV2 HG52 was used to attack Vero cell after transfection overnight.Vero cell and supernatant were collected at 1d,2d,3d,4d and 5d after virus attacking.Flurogenic quantitative reverse transcription polymerase chain reaction(FQ-RT-PCR)was used to detect the expression of HSV2 ICP4 mRNA,flurogenic quantitative polymerase chain reaction (FG-PCR) was used to detect the expression of HSV2 DNA and Western-Blot was used to detect the expression of HSV2 ICP4 protein.Results All the four pairs of siRNA could significantly inhibit the expression of HSV2 ICP4 mRNA and protein,especially siRNA-2.The above siRNAs could significantly decrease HSV2 DNA copy number,too.Conclusion siRNAs targeted to HSV2 ICP4 gene could significantly inhibit expression of HSV2 ICP4 mRNA and protein,and decrease HSV2 DNA copy number,suggesting that siRNA can inhibit HSV2 DNA replication through silencing ICP4 gene.