[Preliminary study of DNA polymerase beta gene silencing by small interfering RNA in human gastric cancer BGC-823 cells].
OBJECTIVE
To study the influence of DNA polymerase beta (polbeta) gene silencing by small interfering RNA on biological behavior of human gastric cancer cell line BGC-823.


METHODS
The siRNA eukaryotic expression vectors targeting polbeta gene were constructed and transfected into BGC-823 cells by liposome. Stable cell lines were screened with G418. The expression levels of polbeta mRNA and protein were detected by real time PCR and Western blot in the cells of each group. The proliferation of each group was detected by flow cytometry and tumorigenicity was determined in nude mice.


RESULTS
The siRNA expression vector targeting polbeta gene was successfully constructed. The expression levels of polbeta mRNA and protein were significantly reduced in the experimental group transfected with siRNA expression vectors targeting polbeta, and the silencing effect of pRNAT-U6.1-sipolbeta2 (suppression degree was 83%) was stronger than that of pRNAT-U6.1-sipolbeta1 (depression degree is 56%). Compared with irrelevant siRNA control group, empty vector control group and untransfected group, the ratio of G0/G1 cells was increased, proportion of S phase cells and cell proliferation were decreased in the experimental group 1 cells transfected with pRNAT-U6.1-sipolbeta1 (P < 0.05). On the contrary, the ratio of G1/G0 was decreased, proportion of S phase cells and cell proliferation was increased in the experimental group 2 cells transfected with pRNAT-U6.1-sipolbeta2 (P < 0.05).


CONCLUSIONS
The siRNA expression vectors targeting DNA polymerase beta gene can significantly inhibit the expression of polbeta mRNA. Neither high nor extremely low expression of polbeta is beneficial to maintain the cellular physiological functions. The expression of polbeta silenced to a proper level by siRNA may play an important role in inhibiting tumorigenesis.
Zhao
Guo-qiang
GQ
Department of Microbiology and Immunology, School of Basic Medical Sciences, Zhengzhou University, Zhengzhou 450001, China.
Wang
Lei
L
Dong
Zi-ming
ZM
D000818:Animals; D002453:Cell Cycle; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D019951:DNA Polymerase beta / Q000235:genetics* / Q000378:metabolism; D020868:Gene Silencing; D005822:Genetic Vectors; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D009368:Neoplasm Transplantation; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering; D011897:Random Allocation; D013274:Stomach Neoplasms / Q000378:metabolism / Q000473:pathology*; D014162:Transfection; D047368:Tumor Burden
D012333:RNA, Messenger; D034741:RNA, Small Interfering; D019951:DNA Polymerase beta