Objective     To construct RNA interference recombinant to inhibit Pygo2 expression and explore its effect on proliferation of glioblastoma U251 cells.      Methods     A pair of specific short hairpin RNA (shRNA) and scrambled (scr) control shRNA were designed accord

ing to Pygo2 cDNA sequence

and then cloned into eukaryotic expression vector pSuper to construct recombinants. After

Eco

RⅠ and

Hin

dⅢ digestion identification and DNA sequencing

the recombinants were used to transfect glioblastoma U251 cells with lipofectamine

TM

2000. RT-PCR and Western blotting were applied to detect the RNA interference effects of Pyro2 shRNA. Colony-forming assay and MTT assay

flow cytometry

and BrdU incorporation assay were employed to detect cell proliferation

cell cycle

and DNA synthesis

respectively.     Results     Double digestion and sequencing results verified that the inserted sequences were correct. Pygo2 shRNA remarkably inhibited the mRNA and protein expression of Pygo2 as well as the proliferation of U251 cells. Comparing with the colony formation rate in the scr shRNA group [(78.9±6.8)%

]

that in the Pygo2 shRNA group [(55.4±5.2)%

]

was significantly reduced (

P

<

0.05). The BrdU incorporation rate in the control group [(20.96±2.19)%

]

was significantly higher than that in the Pygo2 shRNA group [(8.81±0.56)%

]

(

P<

0.05)

but it was similar to that in the scr shRNA group [(20.35±1.73)%

]

. Pygo2 shRNA decreased the U251 cell percentage in S phase and increased that in G1 phase obviously (

P

<

0.01)

inducing cell cycle arrest in G1 phase.     Conclusion     Down-regulation of Pygo2 expression can effectively inhibit DNA synthesis of U251 cells

induce cell cycle arrest in G1 phase

and inhibit cell proliferation.