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  • Title: [Efficient expression of multidrug resistance gene mdr1 in retroviral vector under control of an internal ribosome entry site].
    Author: Fu J, Chen Z, Wang W.
    Journal: Zhonghua Xue Ye Xue Za Zhi; 1998 Dec; 19(12):619-22. PubMed ID: 11263327.
    Abstract:
    OBJECTIVE: To test the efficiency of human mdr1 gene expression under the control of an internal ribosome entry site (IRES). METHODS: The expression retroviral vector Halmdr1 was constructed from pSXLC/pHa system which contains an IRES from encephalomyocarditis virus. The vector was introduced into ecotropic packaging cells GP + E86 by liposome-mediated transfection. The retrovirus producing cells were obtained by 50 micrograms/L vincristine selection. The success of transfer and expression of mdr1 gene were determined by polymerase chain reaction (PCR) and flow cytometry (FCM). RESULTS: Virus in the supernatant of the producer cells, in which the integration of mdr1 gene was confirmed by PCR, was titrated to 2.0 x 10(5) cfu/ml. The selected producer cells exhibited a 24-52-fold increase of resistance to vincristine, daunorubicin and taxol in comparison with control cells GP + E86. The expression of mdr1 gene was confirmed by both reverse transcription PCR at RNA level and FCM at protein level, although the HaImdr1 transfectants showed somewhat less expression of P-gp when compared to that with cap-dependent translation of Ha mdr1. CONCLUSION: mdr1 gene can be effectively translated and expressed under the control of IRES in cap-independent manner, it may be used as a dominant selectable marker in bicistronic vectors for gene therapy.
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