[ShRNA against NSP4 gene inhibits the proliferation of bovine rotavirus in vitro].
Based on the NSP4 sequence of bovine rotavirus (BRV), the shRNA was designed and synthesized, and a shRNA recombinant lenti-virus vector RNAi-H1-89 was constructed. The recombinant RNAi-H1-89 Lenti-virus was packaged by transfecting the 293T cell with the recombinant vector RNAi-H1-89 and two helper plasmids using lipofectamine, and then used to infect MA104 cells. The MA104 cells were further infected with BRV strain G6 24h post-infection, with the LacZ shRNA recombinant lenti-virus as control. Thirty-six hours later, the CPE of the infected cells was observed under microscope, shRNA of NSP4 gene inhibited CPE in MA104 cell; the shRNA against NSP4 gene also inhibited NSP4 gene expression by RT-PCR, The virus titer in the cell culture supernatant was significant lower compared with the control group. The above results showed that RNAi-H1-89 against NSP4 gene could specifically silence NSP4 gene expression, and inhibit the proliferation of BRV.
Chen
Fang-yuan
FY
Life College of Northeast Agricultural University, Harbin 150030, China. chenfangyuan.1983@163.com
Wang
Hong-mei
HM
Yang
Shao-hua
SH
Wu
Jian-ming
JM
Gao
Yun-dong
YD
Liu
Xiao
X
Yang
Hong-jun
HJ
Wang
Chang-fa
CF
Zhong
Ji-feng
JF
Wang
Li-qun
LQ
He
Hong-bin
HB
D000818:Animals; D001483:Base Sequence; D002417:Cattle; D002460:Cell Line; D004274:DNA, Recombinant / Q000235:genetics; D006023:Glycoproteins / Q000172:deficiency* / Q000235:genetics*; D008969:Molecular Sequence Data; D010957:Plasmids / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics*; D012401:Rotavirus / Q000235:genetics* / Q000502:physiology*; D014118:Toxins, Biological / Q000235:genetics*; D019562:Viral Load / Q000235:genetics; D017361:Viral Nonstructural Proteins / Q000172:deficiency* / Q000235:genetics*; D014779:Virus Replication / Q000235:genetics*
D004274:DNA, Recombinant; D006023:Glycoproteins; C081369:NS28 protein, rotavirus; D034741:RNA, Small Interfering; D014118:Toxins, Biological; D017361:Viral Nonstructural Proteins