Study on gene silence of MDR1 gene triggered by small interfering RNA in multidrug resistant breast cancer cells

Objective To illustrate the feasibility of gene silence of MDR1 gene triggered by small interfering RNA (siRNA) in drug resistant breast cancer.Methods The experimental subjects included drug resistant human breast cancer cell line MCF7/ADR and human breast cancer cell line MCF7.The siRNA oligonucleotides strand designed previously was inserted into pSilencer3.1-H1 Hygro vector and then the plasmid was transformed into E.coli.After a whole night,the positive clone could be screened on the plate.From the positive clone,the plasmid was obtained,purified,and sequenced to determine whether the ligation between the siRNA insert and the vector was correct.If the sequence was correct,the purified plasmid was transfected into MCF-7/ADR cells.Subsequently,the MCF7/ADR cells were screened by 100 μg/ml hygromycin for 2 weeks and the survival cells were cultured.The expression rate of P-glycoprotein was measured by flow cytometry and real-time relative quantitative PCR.Eventually,there was a drug resistant experiment on the MCF7/ADR with siRNA.Results The expression rate of P-glycoprotein of MCF7/ADR was decreased from 99.8% which represented the untransformed MCF7/ADR cells expression rate to 12.3% which represented the transformed MCF7/ADR cells expression rate.Similarly,the expression rate of green fluorescence protein of LA795/GFP was declined from 74.8% to 10.6% in the positive control.The result of real-time PCR revealed that the threshold cycle value was increased from 25.22 to 30.64 after MCF7/ADR cells were transfected by siRNA.Conclusion siRNA can trigger gene silence of MDR1 gene in human multidrug resistant breast cancer cell line MCF-7/ADR.This successful operation provides evidence to validate that siRNA may become a new medical technique to reverse drug resistance.