[Construction and identification of Kir2ds4 RNAi lentiviral vector].
This study was aimed to construct a lentiviral vector of RNA interfered (RNAi)-kir2ds4 gene. In accordance with study-confirmed effective sequence of siRNA targeting kir2ds4 gene, the complementary DNA containing both sense and antisense oligonuctide of the targeting sequence was designed, synthesized and inserted into pSicoR-GFP vector containing U6 promoter and GFP sequence. The resulting lentiviral vector containing kir2ds4 shRNA was named as LV-sh kir2ds4, and confirmed by PCR and sequencing. 293T cells were co-transfected with lentiviral vector LV-sh kir2ds4 and packaging system. All virus stocks were produced by Lipofectamine 2000-mediated transfection. The titer of virus was tested according to the expression level of GFP. As a result, PCR and DNA sequencing demonstrated that the lentivirus RNAi vector of kir2ds4 was constructed successfully. The titer of virus tested by expression level of GFP was 6 x 10(8) TU/ml. It is concluded that the lentivirus RNAi vector of kir2ds4 has been successfully constructed.
Dou
Li-Ping
LP
Department of Hematology, General Hospital of PLA, Beijing 100853, China.
DA
Wan-Ming
WM
Wang
Chang
C
Kang
Hui-Yuan
HY
Zhao
Yu
Y
Sun
Jing-Fen
JF
Jin
Hai-Jie
HJ
Wang
Quan-Shun
QS
Gao
Chun-Ji
CJ
Yu
Li
L
D001483:Base Sequence; D005822:Genetic Vectors / Q000235:genetics*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D016086:Lentivirus / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics* / Q000378:metabolism; D054340:Receptors, KIR / Q000096:biosynthesis / Q000235:genetics*
C473953:KIR2DS4 protein, human; D034741:RNA, Small Interfering; D054340:Receptors, KIR; D049452:Green Fluorescent Proteins