[Experimental study of K562 cell apoptosis induced by siRNA].
OBJECTIVE
To construct a siRNA expression vector pBCR6 that produces siRNA against bcr/abl mRNA and detect apoptosis rate of K562 cells after pBCR6 transfection.


METHODS
Template sequence for siRNA was designed, synthesized and inserted into an expression vector pSilencer1.0-U6. Restriction analysis and sequencing were performed to verify the pBCR6 vector. Then pBCR6 was transfected into K562 cells by X-tremeGene Q2. pSilencer1.0-U6 was used as the control. At different time point after transfection, apoptosis rate was determined by Tunel and Annexin V+ PI with FCM.


RESULTS
pBCR6 was verified by restriction analysis and sequencing. The apoptosis rate of K562 cells markedly increased at 48 and 72 hour after transfected with pBCR6, and increased in a time-dependent manner [the apoptosis rate of transfected K562 cells was (47.80 +/- 1.63)% at 72 hrs, whereas the control group was (6.67 +/- 0.37)%, P < 0.0001] No prominent change in apoptosis rate was found in the control.


CONCLUSIONS
The siRNA expression vector against bcr/abl mRNA was successfully constructed. The pilot study showed that pBCR6 could effectively induce K562 cells apoptosis. siRNA may be a new tool for molecular target therapy for chronic myelogenous leukemia.
Chen
Bo-bin
BB
Department of Hematology, Huashan Hospital, Fudan University, Shanghai 200040, China.
Fan
Hua-hua
HH
Lin
Guo-wei
GW
Yuan
Zheng-hong
ZH
Lu
Hua-zhong
HZ
Gao
Li
L
Liu
Yan
Y
D017209:Apoptosis; D001483:Base Sequence; D005434:Flow Cytometry; D016044:Fusion Proteins, bcr-abl / Q000235:genetics*; D005822:Genetic Vectors; D006801:Humans; D020287:In Situ Nick-End Labeling; D020014:K562 Cells; D010957:Plasmids / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics*; D014162:Transfection
D012333:RNA, Messenger; D034741:RNA, Small Interfering; D016044:Fusion Proteins, bcr-abl