Short-hairpin RNAs synthesized by T7 phage polymerase do not induce interferon.
RNA interference (RNAi) mediated by small-interfering RNAs (siRNAs) is a highly effective gene-silencing mechanism with great potential for gene-therapeutic applications. siRNA agents also exert non-target-related biological effects and toxicities, including immune-system stimulation. Specifically, siRNA synthesized from the T7 RNA polymerase system triggers a potent induction of type-I interferon (IFN) in a variety of cells. Single-stranded RNA also stimulates innate cytokine responses in mammals. We found that pppGn (n = 2,3) associated with the 5'-end of the short-hairpin RNA (shRNA) from the T7 RNA polymerase system did not induce detectable amounts of IFN. The residual amount of guanine associated with the 5'-end and hairpin structures of the transcript was proportional to the reduction of the IFN response. Here we describe a T7 pppGn (n = 2,3) shRNA synthesis that does not induce the IFN response, and maintains the full efficacy of siRNA.
Gondai
Takuma
T
Department of Life and Environmental Science, High Technology Research Center, Chiba Institute of Technology, 2-17-1 Tsudanuma, Narashino-shi, Chiba 275-0016 Japan.
Yamaguchi
Kazuya
K
Miyano-Kurosaki
Naoko
N
Habu
Yuichirou
Y
Takaku
Hiroshi
H
D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D006147:Guanine / Q000737:chemistry; D015497:HIV-1 / Q000235:genetics / Q000502:physiology; D006367:HeLa Cells; D006801:Humans; D007370:Interferon Type I / Q000096:biosynthesis*; D010710:Phosphates / Q000737:chemistry; D034622:RNA Interference; D022661:RNA, Untranslated / Q000096:biosynthesis / Q000737:chemistry*; D012367:RNA, Viral / Q000737:chemistry; D014764:Viral Proteins / Q000378:metabolism*; D014779:Virus Replication
D007370:Interferon Type I; D010710:Phosphates; D022661:RNA, Untranslated; D012367:RNA, Viral; D014764:Viral Proteins; D006147:Guanine; C062210:bacteriophage T7 RNA polymerase; D012321:DNA-Directed RNA Polymerases