Silencing of Bc1-2 gene by small hairpin RNA inhibits growth of NB4 cells

To investigate the effects of small hairpin RNA(shRNA) targeting Be1-2 on the growth of NB4 cell line. Two of pairs oligonucleotides for short hairpin expression targeting the coding region of Bc1-2 mRNA were designed and chemically synthesized. Annealed oligonucleotides were inserted into Pgenesil-1 vector downstream of U6 promoter to construct RNAi plasmid. Oligonucleotide with a scrambled sequence was used as a negative control. Recombinant expression vector was identified by enzyme cutting and sequencing. Bc1-2 shRNAs were transfected into NB4 cell with Lipofectamine 2000. Western-Blot of Bc1-2 protein expression in NB4 cells was performed after transfection. The inhibition of cell growth was assessed by a MTT assay. Enzyme cutting and sequencing showed that the insertion sequence was correct. Western-Blot assay showed that the expression level of Bc1-2 protein in NB4 cells decreased after Bc1-2 shRNAs treatment. There was no difference in Bc1-2 protein levels between control shRNA and untreated cells. Viable cells at 72 and 96 h after treatment with Bc1-2 shRNAs were less than that after treatment with control shRNAs and untreated NB4 cells, respectively ( P <0.05). Control shRNA had no significant effect on the growth of the cells. Bc1-2 shRNA could effectively inhibit the growth of NB4 cells. Bc1-2 shRNA might be an effective anti-leukemia candidate.