[Detection of IFN Response of Non-Specific Effects on RNAi.].
BACKGROUND
The RNA interference (RNAi) approach is an efficient and widely used method for silencing specific genes and exploring their functions. However, numerous studies have highlighted the non-specific effects of RNAi experiments and the IFN response is one of the most common non-specific effects. In this research, we will study how to detect the IFN response in RNAi experiments.


METHODS
Five different FLJ20420 siRNAs was transfected into human lung adenocarinoma cell line A549 using LipofectamineTM2000. The alternation of the interferon stimulated genes (ISGs) expression profile was analysed by real-time PCR and microarray assay.


RESULTS
(1) Different FLJ20420 siRNA had different effect on silencing FLJ20420 mRNA expression in A549 cells, and FLJsiRNA-1 and -4 had the better effects to knockdown the expression of FLJ20420 (80% and 90% down-regulation, respectively); (2) In analyzed 16 major ISGs, the expression of 14 of these genes was significantly increased in A549-FLJsiRNA-1 cells (P <0.05), while only 2 of these genes had a significant change in A549-FLJ-siRNA-4 cells. These results indicated an IFN response existed in A549-FLJ-siRNA-1 cells. (3) Microarray assay indicated that the expression of 51 ISGs were increased over 2 times in A549-FLJ-siRNA-1 cells, while only the expression of 6 ISGs were over 2-time in A549-FLJ-siRNA-4 cells. These results confirmed that there was an IFN response of non-specific effects of RNAi in A549 cells transfected with FLJ-siRNA-1.


CONCLUSIONS
In RNAi experiments, siRNA can not only silence the target genes, but also may activate IFN response. The analysis of the expression change of the major ISGs by real-time PCR is a useful approach to explore the IFN response in RNAi experiments.
Liang
Yonggang
Y
Department of Thoracic and Caridovascular Surgery, West China Hospital, Sichuan University, Chengdu 610041, China.
Liu
Hongyu
H
Liu
Baoxing
B
Bai
Yun
Y
Wu
Heng
H
Zhou
Qinghua
Q
Chen
Jun
J