[Reversal of leukemia multidrug resistance by sequence-specific short hairpin RNA].
This study was aimed to design and screen short hairpin RNA (shRNA) molecules targeting multidrug resistance gene (mdr1), as well as to investigate the effects of shRNA expression vector on K562/A02 cells. Mdr1-shRNA expression vector was transfected into K562/A02 cells by lipofectamine 2000, and G418 was added to screen and establish the stable expression cell strain. The expressions of mdr1 mRNA and protein were detected by real-time RT-PCR and Western blot respectively. The sensitivity of cells to chemodrugs after interference were tested by CCK8 assay. The function of p-glycoprotein was determined by Rhodamine 123 efflux experiment. The results showed that all of 4 mdr1-shRNA expression vectors could significantly knockdown the expression of p-glycoprotein as compared with control vector, moreover, the vector targeting 508 - 526 sites of mdr1 gene was the best one. It is concluded that the mdr1-shRNA expression vector gained by screening can significantly knockdown the expression of mdr1 gene and reverse leukemia drug resistance, paving the way for the application of RNAi in the following animal experiments.
Du
Wei-Ting
WT
National Key Labortory of Experimental Hematology, Institute of Hematology & Blood Disease Hospital, Chinese Academy of Medical Science & Peking Union Medical College, Tianjin, China.
Liu
Bin
B
Gu
Dong-Sheng
DS
Han
Zhi-Bo
ZB
Liu
Peng-Xia
PX
Xu
Jie
J
Liang
Lu
L
Zhao
Hai-Feng
HF
Lu
Shi-Hong
SH
Yang
Ren-Chi
RC
D018435:ATP Binding Cassette Transporter, Subfamily B; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000235:genetics*; D001483:Base Sequence; D018432:Drug Resistance, Multiple / Q000235:genetics*; D019008:Drug Resistance, Neoplasm / Q000235:genetics*; D055785:Gene Knockdown Techniques; D019450:Genes, MDR; D005822:Genetic Vectors; D006801:Humans; D020014:K562 Cells; D007938:Leukemia / Q000235:genetics; D034622:RNA Interference; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics*; D014162:Transfection
C513055:ABCB1 protein, human; D018435:ATP Binding Cassette Transporter, Subfamily B; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D012333:RNA, Messenger; D034741:RNA, Small Interfering