کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
2078912 1545052 2008 5 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Construction of Recombinant Retroviral Vector Carrying Lab Gene of Foot-and-mouth Disease Virus and Its Expression in Bovine Kidney (MDBK) Cells
موضوعات مرتبط
علوم زیستی و بیوفناوری بیوشیمی، ژنتیک و زیست شناسی مولکولی بیوتکنولوژی یا زیست‌فناوری
پیش نمایش صفحه اول مقاله
Construction of Recombinant Retroviral Vector Carrying Lab Gene of Foot-and-mouth Disease Virus and Its Expression in Bovine Kidney (MDBK) Cells
چکیده انگلیسی
In this study, foot-and-mouth disease virus (FMDV) strain OA/58 RNAs were used as templates for RT-PCR. By the molecular cloning, the Lab gene encoding leader protease called Lpro were cloned in retroviral vector pBPSTR1 to obtain reconstruction retroviral vector termed pBPSTR1-Lab. At different concentrations of puromycin and tetracycline respectively in the cell culture mediums, the growth of bovine kidney cells (MDBK) showed that the optimal puromycin resistant selection concentration was 3 μg/mL and tetracycline regulatory concentration was 1 μg/mL. Pseudotyped retroviral virus particles were produced by transiently co-transfecting GP2-293 cells with a retroviral vector DNA and VSV-G plasmid. Then MDBK cells were infected by pseudotyped retroviral virus and were continually seeded in the medium at the optimal tetracycline regulatory concentration and puromycin selection concentration for 12 days to obtain puromycin resistant colonies whose genomes contained the Lab gene. After tetracycline removal, synthesis of Lpro induced severe morphological changes in the puromycin resistant MDBK cells. PCR and Western blotting proved that a stable MDBK cell line inducibly expressing the Lab gene under the control of tetracycline was obtained. The experiment might provide a basis for studying that Lpro of FMDV plays an important role in MDBK cell pathogenesis.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Chinese Journal of Biotechnology - Volume 24, Issue 5, May 2008, Pages 740-745
نویسندگان
, , , , , , , ,