[Establishment of a stable nasopharyngeal carcinoma cell line with lentivirus-mediated RNA interference for EIF4G1 gene silencing].
OBJECTIVE
To establish a nasopharyngeal carcinoma (NPC) cell line with stable EIF4G1 gene silencing induced by small interfering RNA (siRNA).


METHODS
The EIF4G1 mRNA levels in 8 NPC cell lines including 5-8F, 6-10B, C666-1, CNE1, CNE2, HNE1, HONE1, and SUNE1 were detected by fluorescence quantitative RT-PCR (QRT-PCR). The recombinant lentivirus shRNA expression plasmid targeting EIF4G1 gene was packaged into mature lentivirus by 293FT cells and used to infect 5-8F cells. After blasticidin selection of NPC cells with constant expression of the EIF4G1-siRNA, the efficiency of EIF4G1 mRNA expression interference was determined using QRT-PCR.


RESULTS
The 8 NPC cell lines showed differential expression of EIF4G1 mRNA, among which 5-8F cells had the highest EIF4G1 expression. The recombinant lentivirus plasmid pLenti6/BLOCK-iT-DEST/EIF4G1-shRNA was successfully constructed and verified by PCR and sequencing. The EIF4G1 mRNA level of 5-8F cells infected with shRNA-EIF4G1 lentivirus was significantly reduced as compared with the negative control and the blank control cells.


CONCLUSIONS
The recombinant lentivirus vector pLenti6/BLOCK- iT-DEST/EIF4G1-shRNA we constructed results in marked downregulation of EIF4G1 mRNA expression and constant expression of EIF4G1-siRNA after infection of 5-8F cells.
Tu
Lu-xia
LX
Cancer Research Institute, Key Lab for Transcriptomics and Proteomics of Human Major Diseases, Southern Medical University, Guangzhou 510515, China.
Fang
Wei-yi
WY
Liu
Zhen
Z
Li
Xin
X
He
Ying
Y
Xie
Si-ming
SM
Yao
Kai-tai
KT
D045744:Cell Line, Tumor; D039603:Eukaryotic Initiation Factor-4G / Q000096:biosynthesis / Q000235:genetics*; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D016086:Lentivirus / Q000235:genetics* / Q000378:metabolism; D009303:Nasopharyngeal Neoplasms / Q000235:genetics* / Q000378:metabolism; D034622:RNA Interference; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics*; D014162:Transfection
C113920:EIF4G1 protein, human; D039603:Eukaryotic Initiation Factor-4G; D012333:RNA, Messenger; D034741:RNA, Small Interfering