[Effect of small interfering RNA targeting Rac1 gene on colony formation of SW480 cells in vitro].
OBJECTIVE
To construct a vector expressing small interfering RNA (siRNA) against Rac1 gene and observe its effect on soft agar colony formation of SW480 cells in vitro.


METHODS
Oligos of 64 base pairs for hairpin RNA targeting Rac1 were chemically synthesized and annealed. The siRNA constructs for Rac1, produced by inserting the annealed oligos into the downstream of H1 promoter of linearized pSUPER, were confirmed by restriction digestion and DNA sequencing. The constructed Rac1-siRNA was transfected into SW480 cells and Western blotting was performed to assess the expression and interference efficiency of siRNAs against Rac1.The soft agar colony formation assay was used to study the effect of Rac1 gene silencing on SW480 cells.


RESULTS
Restriction digestion and DNA sequencing showed that the siRNA targeting Rac1 gene was successfully constructed. The siRNA could effectively down-regulate the expression of Rac1 in SW480 cells. Soft agar colony formation assay showed that the colony number and diameter of SW480 cells was reduced after siRNA transfection.


CONCLUSIONS
A vector expressing hairpin RNA against Rac1 gene are successfully produced, which significantly reduces the colony numbers and size of SW480 cells in vitro, suggesting that Rac1 plays an important role in the growth of colorectal cancer in vitro.
Nan
Qing-zhen
QZ
Institute of Digestive Diseases, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China. nanqingzhen@126.com
Gao
Lei
L
Xiao
Bing
B
Zhang
Zhen-shu
ZS
Jiang
Bo
B
D001483:Base Sequence; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D003110:Colonic Neoplasms / Q000473:pathology*; D015536:Down-Regulation; D006801:Humans; D008969:Molecular Sequence Data; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics*; D014162:Transfection; D020830:rac1 GTP-Binding Protein / Q000235:genetics*
C107953:RAC1 protein, human; D034741:RNA, Small Interfering; D020830:rac1 GTP-Binding Protein