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Cell Journal، جلد ۲۰، شماره ۱، صفحات ۹۸-۱۰۷
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عنوان انگلیسی PhiC31-based Site-Specific Transgenesis System for Production of Transgenic Bovine Embryos by Somatic Cell Nuclear Transfer and Intracytoplasmic Sperm Injection
چکیده انگلیسی مقاله Objective: The Streptomyces phage phiC31 integrase offers a sequence-specific method of transgenesis with a robust long-term gene expression. PhiC31 has been successfully developed in a variety of tissues and organs for purpose of in vivo gene therapy. Materials and methods: In this study, the application of phiC31 integrase system was evaluated for generating transgenic bovine embryos by somatic cell nuclear transfer (SCNT) and sperm mediated gene transfer (SMGT) approaches. Results: PhiC31 integrase mRNA and protein was produced in vitro and their functionality was confirmed. Seven phiC31 recognizable bovine pseudo attP sites were considered for evaluation of site specific recombination. The accuracy of these sites was validated in phic31 targeted bovine fibroblasts using PCR and sequencing. The efficiency and site-specificity of phiC31 integrase system was also confirmed in generated transgenic bovine embryo which successfully obtained using SCNT and SMGT technique. Conclusion: The results showed that both SMGT and SCNT -derived embryos were EGFP positive and phiC31 integrase could recombine the reporter gene in a site specific manner. These results demonstrate that attP site can be used as a proper location to conduct site directed transgenesis in both mammalian cells and embryos in phiC31 integrase system when even combinaed to SCNT and intracytoplasmic sperm injection (ICSI) method.
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نویسندگان مقاله | Mohammad Hadi Sekhavati


| Sayed Morteza Hosseini


| Mojtaba Tahmoorespur


| Kamran Ghaedi


| Farnoosh Jafarpour


| Mehdi Hajian


| Kyanoosh Dormiani


| Mohammad Hossein Nasr Esfahani


نشانی اینترنتی http://celljournal.org/journal/article/abstract/4385
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