We have previously reported that the 2-amino-6-vinylpurine (AVP) in oligonucleotide (ODN) showed the highly selective and effective cross-linking reaction toward cytosine within target complementary sequences. Recently, we revealed that the PEGylated ODN containing AVP analogues exhibited the effective antisense effect in the cultured cell. However, subsequent studies showed that the cross-linking ability of AVP for cytosine was affected by the target RNA sequences. We therefore developed new intelligent nucleoside analogues connected AVP to sugar through the ethylene linker (et-AVP), that exhibited highly effective cross-linking ability to rC in the target RNA. In this paper, we described the synthesis of the PEGylated ODN containing cross-linking nucleoside analogues and properties of it about RNase H activity and antisense effect in cell lysate. As a result, the PEGylated ODN containing et-AVP showed the effective antisense effect in the non-cell system for targeting the luciferase mRNA by non-RNase-H mechanism.